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Cell_Biology_Alberts. Figure 15–63 Negative feedback in the NFκb signaling pathway induces oscillations in NFκb activation. (A) Drawing showing how activated NFκB stimulates the transcription of the IκBα gene, the protein product of which acts back in the cytoplasm to sequester and inhibit NFκB there; if the stimulus is persistent, the newly made IκBα protein will then be ubiquitylated and degraded, liberating active NFκB again so that it can return to the nucleus and activate transcription (see Figure 15–62). (B) A short exposure to TNFα produces a single, short pulse of NFκB activation, beginning within minutes and ending by 1 hour. This response turns on the transcription of gene A but not gene B. (C) A sustained exposure to TNFα for the entire 6 hours of the experiment produces oscillations in NFκB activation that damp down over time. This response turns on the transcription of both genes; gene B turns on only after several hours, indicating that gene B transcription requires prolonged activation of


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Cell_Biology_Alberts. that damp down over time. This response turns on the transcription of both genes; gene B turns on only after several hours, indicating that gene B transcription requires prolonged activation of NFκB, for reasons that are not understood. (D) These time-lapse confocal fluorescence micrographs from a different study of TNFα stimulation show the oscillations of NFκB in a cultured cell, as indicated by its periodic movement into the nucleus (N) of a fusion protein composed of NFκB fused to a red fluorescent protein. In the cell at the center of the micrographs, NFκB is active and in the nucleus at 6, 60, 210, 380, and 480 minutes, but it is exclusively in the cytoplasm at 0, 120, 300, 410, and 510 minutes. (A–C, based on data from A. Hoffmann et al., Science 298:1241–1245, 2002, and adapted from A.Y. Ting and D. Endy, Science 298:1189–1190, 2002; D, from D.E. Nelson et al., Science 306:704–708, 2004. All with permission from AAAS.) chemical structure (Figure 15–64) and function, they all


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Cell_Biology_Alberts. A.Y. Ting and D. Endy, Science 298:1189–1190, 2002; D, from D.E. Nelson et al., Science 306:704–708, 2004. All with permission from AAAS.) chemical structure (Figure 15–64) and function, they all act by a similar mechanism. They bind to their respective intracellular receptor proteins and alter the ability of these proteins to control the transcription of specific genes. Thus, these proteins serve both as intracellular receptors and as intracellular effectors for the signal.


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Cell_Biology_Alberts. The receptors are all structurally related, being part of the very large nuclear receptor superfamily. Many family members have been identified by DNA sequencing only, and their ligand is not yet known; they are therefore referred to as orphan nuclear receptors, and they make up large fractions of the nuclear receptors encoded in the genomes of humans, Drosophila, and the nematode C. elegans. Some mammalian nuclear receptors are regulated by intracellular metabolites rather than by secreted signal molecules; the peroxisome proliferation-activated receptors (PPARs), for example, bind intracellular lipid metabolites and regulate the transcription of genes involved in lipid metabolism and fat-cell differentiation. It seems likely that the nuclear receptors for hormones evolved from such receptors for intracellular metabolites, which would help explain their intracellular location.


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Cell_Biology_Alberts. Steroid hormones—which include cortisol, the steroid sex hormones, vitamin D (in vertebrates), and the molting hormone ecdysone (in insects)—are all made from cholesterol. Cortisol is produced in the cortex of the adrenal glands and influences the metabolism of many types of cells. The steroid sex hormones are made in the testes and ovaries and are responsible for the secondary sex characteristics that distinguish males from females. Vitamin D is synthesized in the skin in response to sunlight; after it has been converted to its active form in the liver or kidneys, it regulates Ca2+ metabolism, promoting Ca2+ uptake in the gut and reducing its excretion in the kidneys. The thyroid hormones, which are made from the amino acid tyrosine, act to increase the metabolic rate of many cell types, while the retinoids, such as retinoic acid, are made from vitamin A and have important roles as local mediators in vertebrate development. Although all of these signal molecules are relatively


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Cell_Biology_Alberts. types, while the retinoids, such as retinoic acid, are made from vitamin A and have important roles as local mediators in vertebrate development. Although all of these signal molecules are relatively insoluble in water, they are made soluble for transport in the bloodstream and other extracellular fluids by binding to specific carrier proteins, from which they dissociate before entering a target cell (see Figure 15–3B).


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Cell_Biology_Alberts. The nuclear receptors bind to specific DNA sequences adjacent to the genes that the ligand regulates. Some of the receptors, such as those for cortisol, are located primarily in the cytosol and enter the nucleus only after ligand binding; others, such as the thyroid and retinoid receptors, are bound to DNA in the nucleus even in the absence of ligand. In either case, the inactive receptors are usually bound to inhibitory protein complexes. Ligand binding alters the conformation of the receptor protein, causing the inhibitory complex to dissociate, while also causing the receptor to bind coactivator proteins that stimulate gene transcription (Figure 15–65). In other cases, however, ligand binding to a nuclear receptor inhibits transcription: some thyroid hormone receptors, for example, act as transcriptional activators in the absence of their hormone and become transcriptional repressors when hormone binds.


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Cell_Biology_Alberts. Thus far, we have focused on the control of gene expression by extracellular signal molecules produced by other cells. We now turn to gene regulation by a more global environmental signal: the cycle of light and darkness that results from the Earth’s rotation. Life on Earth evolved in the presence of a daily cycle of day and night, and many present-day organisms (ranging from archaea to plants and humans) possess an internal rhythm that dictates different behaviors at different times of day. These behaviors range from the cyclical change in metabolic enzyme activities of a bacterium to the elaborate sleep–wake cycles of humans. The internal oscillators that control such diurnal rhythms are called circadian clocks.


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Cell_Biology_Alberts. Having a circadian clock enables an organism to anticipate the regular daily changes in its environment and take appropriate action in advance. Of course, the internal clock cannot be perfectly accurate, and so it must be capable of being reset by external cues such as the light of day. Thus, circadian clocks keep running Figure 15–64 Some signal molecules that bind to intracellular receptors. Note that all of them are small and hydrophobic. The active, hydroxylated form of vitamin D3 is shown. Estradiol and testosterone are steroid sex hormones.


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Cell_Biology_Alberts. ligand ˜ helix of coactivator even when the environmental cues (changes in light and dark) are removed, but the period of this free-running rhythm is generally a little less or more than 24 hours. External signals indicating the time of day cause small adjustments in the running of the clock, so as to keep the organism in synchrony with its environment. Following more drastic shifts, circadian cycles become gradually reset (entrained) by the new cycle of light and dark, as anyone who has experienced jet lag can attest.


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Cell_Biology_Alberts. We might expect that the circadian clock would be a complex multicellular device, with different groups of cells responsible for different parts of the oscillation mechanism. Remarkably, however, in almost all multicellular organisms, including humans, the timekeepers are individual cells. Thus, a clock that operates in each member of a specialized group of brain cells (the SCN cells in the suprachiasmatic nucleus of the hypothalamus) controls our diurnal cycles of sleeping and waking, body temperature, and hormone release. Even if these cells are removed from the brain and dispersed in a culture dish, they will continue to oscillate individually, showing a cyclic pattern of gene expression with a period of approximately 24 hours. In the intact body, the SCN cells receive neural cues from the retina, entraining the SCN cells to the daily cycle of light and dark; they also send information about the time of day to another brain area, the pineal gland, which relays the time signal to


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Cell_Biology_Alberts. the retina, entraining the SCN cells to the daily cycle of light and dark; they also send information about the time of day to another brain area, the pineal gland, which relays the time signal to the rest of the body by releasing the hormone melatonin in time with the clock.


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Cell_Biology_Alberts. Although the SCN cells have a central role as timekeepers in mammals, almost all the other cells in the mammalian body have an internal circadian rhythm, which has the ability to reset in response to light. Similarly, in Drosophila, many different types of cells have a similar circadian clock, which continues to cycle when they have been dissected away from the rest of the fly and can be reset by externally imposed light and dark cycles. The working of circadian clocks, therefore, is a fundamental problem in cell biology. Although we do not yet understand all the details, studies in a wide variety of organisms have revealed the basic principles and molecular components. The key principle is that circadian clocks generally depend on negative feedback loops. As discussed earlier, oscillations in the activity of an intracellular signaling protein can occur if that protein inhibits its own activity with a long delay (see


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Cell_Biology_Alberts. Figure 15–65 The activation of nuclear receptors. All nuclear receptors bind to DNA as either homodimers or heterodimers, but for simplicity we show them as monomers. (A) The receptors all have a related structure, which includes three major domains, as shown. An inactive receptor is bound to inhibitory proteins. Typically, the binding of ligand to the receptor causes the ligand-binding domain of the receptor to clamp shut around the ligand, the inhibitory proteins to dissociate, and coactivator proteins to bind to the receptor’s transcription-activating domain, thereby increasing gene transcription. In other cases, ligand binding has the opposite effect, causing co-repressor proteins to bind to the receptor, thereby decreasing transcription (not shown).


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Cell_Biology_Alberts. The structure of the ligand-binding domain of the retinoic acid receptor is shown in the absence (left) and presence (middle) of ligand (shown in red). When ligand binds, the blue α helix acts as a lid that snaps shut, trapping the ligand in place. The shift in the conformation of the receptor upon ligand binding also creates a binding site for a small α helix (orange) on the surface of coactivator proteins. (PDB codes: 1LBD, 2ZYO, and 2ZXZ.) Tim degraded in response to light mRNAs dissociation of heterodimer


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Cell_Biology_Alberts. Tim degraded in response to light mRNAs dissociation of heterodimer Figure 15–18C and D). In Drosophila and many other animals, including humans, the heart of the circadian clock is a delayed negative feedback loop based on transcription regulators: accumulation of certain gene products switches off the transcription of their own genes, but with a delay, so that the cell oscillates between a state in which the products are present and transcription is switched off, and one in which the products are absent and transcription is switched on (Figure 15–66). The negative feedback underlying circadian rhythms does not have to be based on transcription regulators. In some cell types, the circadian clock is constructed of proteins that govern their own activities through post-translational mechanisms, as we discuss next.


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Cell_Biology_Alberts. The best understood circadian clock is found in the photosynthetic cyanobacterium, Synechococcus elongatus. The core oscillator in this organism is remarkably simple, being composed of just three proteins—KaiA, KaiB, and KaiC. The central player is KaiC, a multifunctional enzyme that catalyzes its own phosphorylation and dephosphorylation in a 24-hour cycle: it gradually phosphorylates itself sequentially at two sites during the day and dephosphorylates itself during the night. This timing depends on interactions with the two other Kai proteins: KaiA binds to unphosphorylated KaiC and stimulates KaiC autophosphosphorylation, first at one site and then, with a delay, at the other. The second phosphorylation promotes the binding of the third protein, KaiB, which blocks the stimulatory effect of KaiA and thereby allows KaiC to dephosphorylate itself, bringing KaiC back to its dephosphorylated state. This clock depends on a negative feedback loop: KaiC drives its own phosphorylation


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Cell_Biology_Alberts. effect of KaiA and thereby allows KaiC to dephosphorylate itself, bringing KaiC back to its dephosphorylated state. This clock depends on a negative feedback loop: KaiC drives its own phosphorylation until, after a delay, it recruits an inhibitor, KaiB, that stimulates KaiC to dephosphorylate itself. Amazingly, when the three Kai proteins are purified and incubated in a test tube with ATP, KaiC phosphorylation and dephosphorylation occur with roughly 24-hour timing over a period of several days (Figure 15–67).


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Cell_Biology_Alberts. Circadian oscillations in KaiC phosphorylation lead to parallel rhythms in the expression of large numbers of genes involved in controlling metabolic activities and cell division (see Figure 15–67). As a result, many aspects of cell behavior are synchronized with the circadian cycle. Even in continuous darkness, cyanobacterial cells generate free-running oscillations of KaiC phosphorylation with roughly 24-hour periods. As in other circadian clocks, the cyanobacterial clock is entrained by the environmental light/dark


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Cell_Biology_Alberts. Figure 15–66 Simplified outline of the mechanism of the circadian clock in Drosophila cells. A central feature of the clock is the periodic accumulation and decay of two transcription regulatory proteins, Tim (short for timeless, based on the phenotype of a gene mutation) and Per (short for period). The mRNAs encoding these proteins rise gradually during the day and are translated in the cytosol, where the two proteins associate to form a heterodimer. After a time delay, the heterodimer dissociates and Tim and Per are transported into the nucleus, where Per represses the Tim and Per genes, resulting in negative feedback that causes the levels of Tim and Per to fall. In addition to this transcriptional feedback, the clock depends on numerous other proteins. For example, the controlled degradation of Per indicated in the diagram imposes delays in the accumulation of Tim and Per, which are crucial to the functioning of the clock. Steps at which specific delays are imposed are shown in


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Cell_Biology_Alberts. of Per indicated in the diagram imposes delays in the accumulation of Tim and Per, which are crucial to the functioning of the clock. Steps at which specific delays are imposed are shown in red.


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Cell_Biology_Alberts. Entrainment (or resetting) of the clock occurs in response to new light–dark cycles. Although most Drosophila cells do not have true photoreceptors, light is sensed by intracellular flavoproteins, also called cryptochromes. In the presence of light, these proteins associate with the Tim protein and cause its degradation, thereby resetting the clock. (Adapted from J.C. Dunlap, Science 311:184–186, 2006.) 1.2 1.0 0.8 0.6 0.4 0.2 cycle. Light is thought to affect the circadian clock indirectly: the activities of Kai proteins are influenced by changes in intracellular redox potential, which occur as a result of increased photosynthetic activity during the day.


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Cell_Biology_Alberts. Some signaling pathways that are especially important in animal development depend on proteolysis to control the activity and location of latent transcription regulatory proteins. Notch receptors are themselves such proteins, which are activated by cleavage when Delta on another cell binds to them; the cleaved cytosolic tail of Notch migrates into the nucleus, where it stimulates the transcription of Notch-responsive genes. In the Wnt/β-catenin signaling pathway, by contrast, the proteolysis of the latent transcription regulatory protein β-catenin is inhibited when a secreted Wnt protein binds to both a Frizzled and LRP receptor protein; as a result, β-catenin accumulates in the nucleus and activates the transcription of Wnt target genes.


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Cell_Biology_Alberts. Hedgehog signaling in flies works much like Wnt signaling. In the absence of a signal, a bifunctional, cytoplasmic transcription regulator, Ci, is proteolytically cleaved to form a transcriptional repressor that keeps Hedgehog target genes silenced. The binding of Hedgehog to its receptors (Patched and iHog) inhibits the proteolytic processing of Ci; as a result, the intact Ci protein accumulates in the nucleus and activates the transcription of Hedgehog-responsive genes. In Notch, Wnt, and Hedgehog signaling, the extracellular signal triggers a switch from transcriptional repression to transcriptional activation.


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Cell_Biology_Alberts. Signaling through the latent transcription regulator NFκB also depends on proteolysis. NFκB proteins are normally held in an inactive state by inhibitory IκB proteins in the cytoplasm. A variety of extracellular stimuli, including proinflammatory cytokines, trigger the phosphorylation and ubiquitylation of IκB, marking it for degradation; this enables the NFκB to translocate to the nucleus and activate


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Cell_Biology_Alberts. Figure 15–67 The core circadian oscillator of cyanobacteria. (A) KaiC is a combined kinase and phosphatase that phosphorylates and dephosphorylates itself on two adjacent sites. In the absence of other proteins, the phosphatase activity is dominant, and the protein is mostly unphosphorylated. The binding of KaiA to KaiC suppresses the phosphatase activity and promotes the kinase activity, leading to KaiC phosphorylation, first at site 1 and then at site 2, resulting in diphosphorylated KaiC. KaiC then dephosphorylates itself slowly at site 1, even in the presence of KaiA, so that KaiC is phosphorylated only at site 2. This form of KaiC interacts with KaiB, which blocks the stimulatory effects of KaiA, thereby reducing the rate of KaiC phosphorylation and allowing dephosphorylation to occur. Diphosphorylated KaiC increases in abundance during the day and peaks around dusk. It activates other proteins that phosphorylate a transcription regulator (RpaA), which then stimulates expression


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Cell_Biology_Alberts. Diphosphorylated KaiC increases in abundance during the day and peaks around dusk. It activates other proteins that phosphorylate a transcription regulator (RpaA), which then stimulates expression of some genes (the dusk genes that peak in early evening) and inhibits expression of other genes (the dawn genes that peak in the morning). When KaiC dephosphorylation gradually occurs during the night, these effects are reversed: dusk genes are turned off and dawn genes are turned on.


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Cell_Biology_Alberts. (B) In this experiment, the three Kai proteins were purified and mixed in a test tube with ATP (which is required for KaiC kinase activity). Every two hours over the next 3 days, the KaiC protein was analyzed by polyacrylamide gel electrophoresis, in which the phosphorylated form of KaiC migrates more slowly (upper band, P-KaiC) than the nonphosphorylated form (lower band, NP-KaiC). The three different phosphorylated forms of KaiC are not distinguished by this method. The phosphorylation of KaiC oscillates with a roughly 24-hour period. (C) The amount of phosphorylated and unphosphorylated KaiC in the experiment in B is plotted on this graph, along with the amount of total protein. (B and C, from M. Nakajima et al., Science 308:414–415, 2005. With permission from AAAS.) the transcription of its target genes. NFκB also activates the transcription of the gene that encodes IκBα, creating a negative feedback loop, which can produce prolonged oscillations in NFκB activity with sustained


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Cell_Biology_Alberts. of its target genes. NFκB also activates the transcription of the gene that encodes IκBα, creating a negative feedback loop, which can produce prolonged oscillations in NFκB activity with sustained extracellular signaling.


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Cell_Biology_Alberts. Some small, hydrophobic signal molecules, including steroid and thyroid hormones, diffuse across the plasma membrane of the target cell and activate intracellular receptor proteins that directly regulate the transcription of specific genes. In many cell types, gene expression is governed by circadian clocks, in which delayed negative feedback produces 24-hour oscillations in the activities of transcription regulators, anticipating the cell's changing needs during the day and night.


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Cell_Biology_Alberts. In plants, as in animals, cells are in constant communication with one another. Plant cells communicate to coordinate their activities in response to the changing conditions of light, dark, and temperature, which guide the plant’s cycle of growth, flowering, and fruiting. Plant cells also communicate to coordinate activities in their roots, stems, and leaves. In this final section, we consider how plant cells signal to one another and how they respond to light. Less is known about the receptors and intracellular signaling mechanisms involved in cell communication in plants than is known in animals, and we will concentrate mainly on how the receptors and intracellular signaling mechanisms differ from those used by animals.


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Cell_Biology_Alberts. Although plants and animals are both eukaryotes, they have evolved separately for more than a billion years. Their last common ancestor is thought to have been a unicellular eukaryote that had mitochondria but no chloroplasts; the plant lineage acquired chloroplasts after plants and animals diverged. The earliest fossils of multicellular animals and plants date from almost 600 million years ago. Thus, it seems that plants and animals evolved multicellularity independently, each starting from a different unicellular eukaryote, some time between 1.6 and 0.6 billion years ago (Figure 15–68).


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Cell_Biology_Alberts. If multicellularity evolved independently in plants and animals, the molecules and mechanisms used for cell communication will have evolved separately and would be expected to be different. There should be some degree of resemblance, however, because the genes in both plants and animals diverged from those contained by their last common unicellular ancestor. Thus, whereas both plants and animals use nitric oxide, cyclic GMP, Ca2+, and Rho family GTPases for signaling, there are no homologs of the nuclear receptor family, Ras, JAK, STAT, TGFβ, Figure 15–68 The proposed divergence of plant and animal lineages from a common unicellular eukaryotic ancestor. The plant lineage acquired chloroplasts after the two lineages diverged. Both lineages independently gave rise to multicellular organisms—plants and animals. (Paintings courtesy of John Innes Foundation.) unicellular ancestor of animals unicellular ancestor of plants CHLOROPLASTS chloroplast


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Cell_Biology_Alberts. Notch, Wnt, or Hedgehog encoded by the completely sequenced genome of Arabidopsis thaliana, the small flowering plant. Similarly, plants do not seem to use cyclic AMP for intracellular signaling. Nevertheless, the general strategies underlying signaling are frequently very similar in plants and animals. Both, for example, use enzyme-coupled cell-surface receptors, as we now discuss. Receptor Serine/Threonine Kinases Are the Largest Class of Cell-Surface Receptors in Plants


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Cell_Biology_Alberts. Receptor Serine/Threonine Kinases Are the Largest Class of Cell-Surface Receptors in Plants Most cell-surface receptors in plants are enzyme-coupled. However, whereas the largest class of enzyme-coupled receptors in animals is the receptor tyrosine kinase (RTK) class, this type of receptor is extremely rare in plants. Instead, plants rely largely on a great diversity of transmembrane receptor serine/threonine kinases, which have a typical serine/threonine kinase cytoplasmic domain and an extracellular ligand-binding domain. The most abundant types of these receptors have a tandem array of extracellular leucine-rich repeat structures and are therefore called leucine-rich repeat (LRR) receptor kinases.


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Cell_Biology_Alberts. There are about 175 LRR receptor kinases encoded by the Arabidopsis genome. These include a protein called Bri1, which forms part of a cell-surface steroid hormone receptor. Plants synthesize a class of steroids that are called brassinosteroids because they were originally identified in the mustard family Brassicaceae, which includes Arabidopsis. These signal molecules regulate the growth and differentiation of plants throughout their life cycle. Binding of a brassinosteroid to a Bri1 cell-surface receptor kinase initiates an intracellular signaling cascade that uses a GSK3 protein kinase and a protein phosphatase to regulate the phosphorylation and degradation of specific transcription regulatory proteins in the nucleus, and thereby specific gene transcription. Mutant plants that are deficient in the Bri1 receptor kinase are insensitive to brassinosteroids and are therefore dwarfs.


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Cell_Biology_Alberts. The LRR receptor kinases are only one of many classes of transmembrane receptor serine/threonine kinases in plants. There are at least six additional families, each with its own characteristic set of extracellular domains. The lectin receptor kinases, for example, have extracellular domains that bind carbohydrate signal molecules. The Arabidopsis genome encodes over 300 receptor serine/threonine kinases, which makes them the largest family of receptors known in plants. Many are involved in defense responses against pathogens. Ethylene Blocks the Degradation of Specific Transcription Regulatory Proteins in the Nucleus


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Cell_Biology_Alberts. Ethylene Blocks the Degradation of Specific Transcription Regulatory Proteins in the Nucleus Various plant growth regulators (also called plant hormones) help to coordinate plant development. They include ethylene, auxin, cytokinins, gibberellins, and abscisic acid, as well as brassinosteroids. Growth regulators are all small molecules made by most plant cells. They diffuse readily through cell walls and can either act locally or be transported to influence cells further away. Each growth regulator can have multiple effects. The specific effect depends on environmental conditions, the nutritional state of the plant, the responsiveness of the target cells, and which other growth regulators are acting.


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Cell_Biology_Alberts. Ethylene is an important example. This small gas molecule (Figure 15–69A) can influence plant development in various ways; it can, for example, promote fruit ripening, leaf abscission, and plant senescence. It also functions as a stress signal in response to wounding, infection, flooding, and so on. When the shoot of a germinating seedling, for instance, encounters an obstacle, ethylene promotes a complex response that allows the seedling to safely bypass the obstacle (Figure 15–69B and C). Plants have various ethylene receptors, which are located in the endoplasmic reticulum and are all structurally related. They are dimeric, multipass transmembrane proteins, with a copper-containing ethylene-binding domain and a domain that interacts with a cytoplasmic protein called CTR1, which is closely related Figure 15–69 The ethylene-mediated triple response that occurs when the growing shoot of a germinating seedling encounters an obstacle underground.


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Cell_Biology_Alberts. (A) The structure of ethylene. (B) In the absence of obstacles, the shoot grows upward and is long and thin. (C) If the shoot encounters an obstacle, such as a piece of gravel in the soil, the seedling responds to the encounter in three ways. First, it thickens its stem, which can then exert more force on the obstacle. Second, it shields the tip of the shoot (at top) by increasing the curvature of a specialized hook structure. Third, it reduces the shoot’s tendency to grow away from the direction of gravity, so as to avoid the obstacle. (Courtesy of Melanie Webb.) in sequence to the Raf MAP kinase kinase kinase discussed earlier (see Figure 15–49). Surprisingly, it is the empty receptors that are active and keep CTR1 active. By an unknown signaling mechanism, active CTR1 stimulates the ubiquitylation and degradation in proteasomes of a nuclear transcription regulator called EIN3, which is required for the transcription of ethylene-responsive genes. In this way, the empty but active


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Cell_Biology_Alberts. ubiquitylation and degradation in proteasomes of a nuclear transcription regulator called EIN3, which is required for the transcription of ethylene-responsive genes. In this way, the empty but active receptors keep ethylene-response genes off. Ethylene binding inactivates the receptors, altering their conformation so that they no longer activate CTR1. The EIN3 protein is no longer ubiquitylated and degraded and can now activate the transcription of the large number of ethylene-responsive genes (Figure 15–70).


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Cell_Biology_Alberts. Regulated Positioning of Auxin Transporters Patterns Plant Growth The plant hormone auxin, which is generally indole-3-acetic acid (Figure 15–71A), binds to receptor proteins in the nucleus. It helps plants grow toward light, grow upward rather than branch out, and grow their roots downward. It also regulates organ initiation and positioning and helps plants flower and bear fruit. Like ethylene (and like some of the animal signal molecules we have described in this chapter), auxin influences gene expression by controlling the degradation of transcription regulators. It works by stimulating the ubiquitylation and degradation of repressor proteins that block the transcription of auxin target genes in unstimulated cells (Figure 15–71B and C). Auxin is unique in the way that it is transported. Unlike animal hormones, which are usually secreted by a specific endocrine organ and transported to target


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Cell_Biology_Alberts. Auxin is unique in the way that it is transported. Unlike animal hormones, which are usually secreted by a specific endocrine organ and transported to target Figure 15–70 The ethylene signaling pathway. (A) In the absence of ethylene, the receptors and CTR1 are active, causing the ubiquitylation and destruction of EIN3, the transcription regulatory protein in the nucleus that is responsible for the transcription of ethylene-responsive genes. (B) The binding of ethylene inactivates the receptors and disrupts the activation of CTR1. The EIN3 protein is not degraded and can therefore activate the transcription of ethylene-responsive genes.


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Cell_Biology_Alberts. Figure 15–71 The auxin signaling pathway. (A) The structure of the auxin indole-3-acetic acid. (B) In the absence of auxin, a transcriptional repressor protein (called Aux/IAA) binds and suppresses a transcription regulatory protein (called auxin-response factor, ARF), which is required for the transcription of auxin-responsive genes. (C) The auxin receptor proteins are mainly located in the nucleus and form part of ubiquitin ligase complexes (not shown). When activated by auxin binding, the receptor–auxin complexes recruit the ubiquitin ligase ARF complexes, which ubiquitylate the Aux/IAA proteins, marking them for degradation in proteasomes. ARF is now free to activate the transcription of auxin-responsive genes. There are many ARFs, Aux/IAA proteins, and auxin receptors that work as illustrated.


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Cell_Biology_Alberts. cells via the circulatory system, auxin has its own transport system. Specific plasma-membrane-bound influx transporter proteins and efflux transporter proteins move auxin into and out of plant cells, respectively. The efflux transporters can be distributed asymmetrically in the plasma membrane to make the efflux of auxin directional. A row of cells with their auxin efflux transporters confined to the basal plasma membrane, for example, will transport auxin from the top of the plant to the bottom.


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Cell_Biology_Alberts. In some regions of the plant, the localization of the auxin transporters, and therefore the direction of auxin flow, is highly dynamic and regulated. A cell can rapidly redistribute transporters by controlling the traffic of vesicles containing them. The auxin efflux transporters, for example, normally recycle between intracellular vesicles and the plasma membrane. A cell can redistribute these transporters on its surface by inhibiting their endocytosis in one domain of the plasma membrane, causing the transporters to accumulate there. One example occurs in the root, where gravity influences the direction of growth. The auxin efflux transporters are normally distributed symmetrically in the cap cells of the root. Within minutes of a change in the direction of the gravity vector, however, the efflux transporters redistribute to one side of the cells, so that auxin is pumped out toward the side of the root pointing downward. Because auxin inhibits root-cell elongation, this redirection


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Cell_Biology_Alberts. the efflux transporters redistribute to one side of the cells, so that auxin is pumped out toward the side of the root pointing downward. Because auxin inhibits root-cell elongation, this redirection of auxin transport causes the root tip to reorient, so that it grows downward again (Figure 15–72).


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Cell_Biology_Alberts. Phytochromes Detect Red Light, and Cryptochromes Detect Blue Light Plant development is greatly influenced by environmental conditions. Unlike animals, plants cannot move when conditions become unfavorable; they have to adapt or they die. The most important environmental influence on plants is of root downward side of root


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Cell_Biology_Alberts. Figure 15–72 Auxin transport and root gravitropism. (A–C) Roots respond to a 90° change in the gravity vector and adjust their direction of growth so that they grow downward again. The cells that respond to gravity are in the center of the root cap, while it is the epidermal cells further back (on the lower side) that decrease their rate of elongation to restore downward growth. (D) The gravity-responsive cells in the root cap redistribute their auxin efflux transporters in response to the displacement of the root. This redirects the auxin flux mainly to the lower part of the displaced root, where it inhibits the elongation of the epidermal cells. The resulting asymmetrical distribution of auxin in the Arabidopsis root tip shown here is assessed indirectly, using an auxin-responsive reporter gene that encodes a protein fused to green fluorescent protein (GFP); the epidermal cells on the downward side of the root are green, whereas those on the upper side are not, reflecting the


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Cell_Biology_Alberts. reporter gene that encodes a protein fused to green fluorescent protein (GFP); the epidermal cells on the downward side of the root are green, whereas those on the upper side are not, reflecting the asymmetrical distribution of auxin. The distribution of auxin efflux transporters in the plasma membrane of cells in different regions of the root (shown as gray rectangles) is indicated in red, and the direction of auxin efflux is indicated by a green arrow. (The fluorescence photograph in D is from


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Cell_Biology_Alberts. T. Paciorek et al., Nature 435:1251–1256, 2005. With permission from Macmillan Publishers Ltd.) light, which is their energy source and has a major role throughout their entire life cycle—from germination, through seedling development, to flowering and senescence. Plants have thus evolved a large set of light-sensitive proteins to monitor the quantity, quality, direction, and duration of light. These are usually referred to as photoreceptors. However, because the term photoreceptor is also used for light-sensitive cells in the animal retina (see Figure 15–38), we shall use the term photoprotein instead.


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Cell_Biology_Alberts. All photoproteins sense light by means of a covalently attached light-absorbing chromophore, which changes its shape in response to light and then induces a change in the protein’s conformation. The best-known plant photoproteins are the phytochromes, which are present in all plants and in some algae but are absent in animals. These are dimeric, cytoplasmic serine/threonine kinases, which respond differentially and reversibly to red and far-red light: whereas red light usually activates the kinase activity of the phytochrome, far-red light inactivates it. When activated by red light, the phytochrome is thought to phosphorylate itself and then to phosphorylate one or more other proteins in the cell. In some light responses, the activated phytochrome translocates into the nucleus, where it activates transcription regulators to alter gene transcription (Figure 15–73). In other cases, the activated phytochrome activates a latent transcription regulator in the cytoplasm, which then


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Cell_Biology_Alberts. it activates transcription regulators to alter gene transcription (Figure 15–73). In other cases, the activated phytochrome activates a latent transcription regulator in the cytoplasm, which then translocates into the nucleus to regulate gene transcription. In still other cases, the photoprotein triggers signaling pathways in the cytosol that alter the cell’s behavior without involving the nucleus.


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Cell_Biology_Alberts. Plants sense blue light using photoproteins of two other sorts, phototropin and cryptochromes. Phototropin is associated with the plasma membrane and is partly responsible for phototropism, the tendency of plants to grow toward light. Phototropism occurs by directional cell elongation, which is stimulated by auxin, but the links between phototropin and auxin are unknown. Cryptochromes are flavoproteins that are sensitive to blue light. They are structurally related to blue-light-sensitive enzymes called photolyases, which are involved in the repair of ultraviolet-induced DNA damage in all organisms, except most mammals. Unlike phytochromes, cryptochromes are also found in animals, where they have an important role in circadian clocks (see Figure 15–66). Although cryptochromes are thought to have evolved from the photolyases, they do not have a role in DNA repair.


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Cell_Biology_Alberts. Plants and animals are thought to have evolved multicellularity and cell communication mechanisms independently, each starting from a different unicellular eukaryote, which in turn evolved from a common unicellular eukaryotic ancestor. Not surprisingly, therefore, the mechanisms used to signal between cells in animals and in plants have both similarities and differences. Whereas animals rely heavily on GPCRs and RTKs, plants rely mainly on enzyme-coupled receptors of the receptor serine/threonine kinase type, especially ones with extracellular leucinerich repeats. Various plant hormones, or growth regulators, including ethylene and auxin, help coordinate plant development. Ethylene acts through intracellular receptors to stop the degradation of specific nuclear transcription regulators, which can then activate the transcription of ethylene-responsive genes. The receptors for some other plant hormones, including auxin, also regulate the degradation of specific transcription regulators,


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Cell_Biology_Alberts. can then activate the transcription of ethylene-responsive genes. The receptors for some other plant hormones, including auxin, also regulate the degradation of specific transcription regulators, although the details vary. Auxin signaling is unusual in that it has its own highly regulated transport system, in which the dynamic positioning of plasma-membrane-bound auxin transporters controls the direction of auxin flow and thereby the direction of plant growth. Light has an important role in regulating plant development. These light responses are mediated by a variety of light-sensitive photoproteins, including phytochromes, which are responsive to red light, and cryptochromes and phototropin, which are sensitive to blue light.


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Cell_Biology_Alberts. Figure 15–73 One way in which phytochromes mediate a light response in plant cells. When activated by red light, the phytochrome, which is a dimeric protein kinase, phosphorylates itself and then moves into the nucleus, where it activates transcription regulatory proteins to stimulate the transcription of red-lightresponsive genes. How does a cell integrate the information received from its many different cell-surface receptors to make all-or-none decisions? Much of what we know about cell signaling comes from biochemical studies of isolated proteins in test tubes. What is the precise quantitative behavior of intracellular signaling networks in an intact cell, or in an intact animal, where countless other signals and cell components might influence signaling specificity and strength? How do intracellular signaling circuits generate specific and dynamic signaling patterns such as oscillations and waves, and how are these patterns sensed and interpreted by the cell?


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Cell_Biology_Alberts. How do intracellular signaling circuits generate specific and dynamic signaling patterns such as oscillations and waves, and how are these patterns sensed and interpreted by the cell? Scaffold proteins and activated receptor tyrosine kinases nucleate the assembly of large intracellular signaling complexes. What is the dynamic behavior of these complexes, and how does this behavior influence downstream signaling? Which statements are true? explain why or why not. 15–1 All second messengers are water-soluble and diffuse freely through the cytosol. 15–2 In the regulation of molecular switches, protein kinases and guanine nucleotide exchange factors (GEFs) always turn proteins on, whereas protein phosphatases and GTPase-activating proteins (GAPs) always turn proteins off. 15–3 Most intracellular signaling pathways provide numerous opportunities for amplifying the responses to extracellular signals.


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Cell_Biology_Alberts. 15–3 Most intracellular signaling pathways provide numerous opportunities for amplifying the responses to extracellular signals. 15–4 Binding of extracellular ligands to receptor tyrosine kinases (RTKs) activates the intracellular catalytic domain by propagating a conformational change across the lipid bilayer through a single transmembrane α helix. 15–5 Protein tyrosine phosphatases display exquisite specificity for their substrates, unlike most serine/threonine protein phosphatases, which have rather broad specificity. 15–6 Even though plants and animals independently evolved multicellularity, they use virtually all the same signaling proteins and second messengers for cell–cell communication. Discuss the following problems. 15–7 Suppose that the circulating concentration of hormone is 10–10 M and the Kd for binding to its receptor is 10–8


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Cell_Biology_Alberts. Discuss the following problems. 15–7 Suppose that the circulating concentration of hormone is 10–10 M and the Kd for binding to its receptor is 10–8 M. What fraction of the receptors will have hormone bound? If a meaningful physiological response occurs when 50% of the receptors have bound a hormone molecule, how much will the concentration of hormone have to rise to elicit a response? The fraction of receptors (R) bound to hormone (H) to form a receptor–hormone complex (R–H) is [R–H]/ ([R] + [R–H]) = [R–H]/[R]TOT = [H]/([H] + Kd). 15–8 Cells communicate in ways that resemble human communication. Decide which of the following forms of human communication are analogous to autocrine, paracrine, endocrine, and synaptic signaling by cells. A. A telephone conversation b. Talking to people at a cocktail party C. D.


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Cell_Biology_Alberts. A. A telephone conversation b. Talking to people at a cocktail party C. D. Talking to yourself 15–9 Why do signaling responses that involve changes in proteins already present in the cell occur in milliseconds to seconds, whereas responses that require changes in gene expression require minutes to hours? 15–10 How is it that different cells can respond in different ways to exactly the same signaling molecule even when they have identical receptors? 15–11 Why do you suppose that phosphorylation/ dephosphorylation, as opposed to allosteric binding of small molecules, for example, has evolved to play such a prominent role in switching proteins on and off in signaling pathways?


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Cell_Biology_Alberts. 15–12 Consider a signaling pathway that proceeds through three protein kinases that are sequentially activated by phosphorylation. In one case, the kinases are held in a signaling complex by a scaffolding protein; in the other, the kinases are freely diffusible (Figure Q15–1). Discuss the properties of these two types of organization in terms of signal amplification, speed, and potential for cross-talk between signaling pathways. Figure Q15–1 A kinase cascade organized by a scaffolding protein or composed of freely diffusing components (Problem 15–12). 15–13 Describe three ways in which a gradual increase in an extracellular signal can be sharpened by the target cell to produce an abrupt or nearly all-or-none response.


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Cell_Biology_Alberts. 15–14 Activation (“maturation”) of frog oocytes is signaled through a MAP kinase signaling module. An increase in the hormone progesterone triggers the module by stimulating the translation of Mos mRNA, which is the frog’s MAP kinase kinase kinase (Figure Q15–2). Maturation is easy to score visually by the presence of a white spot in the middle of the brown surface of the oocyte (see Figure Q15–2). To determine the dose–response curve for pro-gesterone-induced activation of MAP kinase, you place 16 oocytes in each of six plastic dishes and add various concentrations of progesterone. After an overnight incubation, you crush the oocytes, prepare an extract, and determine the state of MAP kinase phosphorylation (hence, activation) by SDS polyacrylamide-gel electrophoresis (Figure Q15–3A). This analysis shows a graded response of MAP kinase to increasing concentrations of progesterone.


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Cell_Biology_Alberts. progesterone Figure Q15–2 Progesterone-induced MAP kinase activation, leading to oocyte Mos maturation (Problem 15–14). (Courtesy of Helfrid Hochegger.) (A) POOLED OOCYTES –+ of frog oocytes (Problem 15–14). pooled oocytes. (B) Phosphorylation of MAP kinase in individual oocytes. MAP kinase 0.001 0.01 0.1 1 10 immunoblotting using progesterone (µM) a MAP-kinase-specific antibody. The first two lanes in each gel contain nonphosphorylated, –+ 0.03µM progesterone and phosphorylated, active MAP kinase (+). (From J.E. Ferrell, Jr., and 0.1 µM progesterone E.M. Machleder, Science 280:895–898, 1998. 0.3 µM progesterone With permission from AAAS.)


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Cell_Biology_Alberts. E.M. Machleder, Science 280:895–898, 1998. 0.3 µM progesterone With permission from AAAS.) Before you crushed the oocytes, you noticed that not all oocytes in individual dishes had white spots. Had some oocytes undergone partial activation and not yet reached the white-spot stage? To answer this question, you repeat the experiment, but this time you analyze MAP kinase activation in individual oocytes. You are surprised to find that each oocyte has either a fully activated or a completely inactive MAP kinase (Figure Q15–3B). How can an all-ornone response in individual oocytes give rise to a graded response in the population? 15–15 Propose specific types of mutations in the gene for the regulatory subunit of cyclic-AMP-dependent protein kinase (PKA) that could lead to either a permanently active PKA or a permanently inactive PKA.


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Cell_Biology_Alberts. 15–16 Phosphorylase kinase integrates signals from the cyclic-AMP-dependent and Ca2+-dependent signaling pathways that control glycogen breakdown in liver and muscle cells (Figure Q15–4). Phosphorylase kinase is composed of four subunits. One is the protein kinase that catalyzes the addition of phosphate to glycogen phosphorylase to activate it for glycogen breakdown. The other three subunits are regulatory proteins that control the activity of the Figure Q15–4 Integration of cyclic-AMP-dependent and Ca2+dependent signaling pathways by phosphorylase kinase in liver and muscle cells (Problem 15–16).


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Cell_Biology_Alberts. Figure Q15–4 Integration of cyclic-AMP-dependent and Ca2+dependent signaling pathways by phosphorylase kinase in liver and muscle cells (Problem 15–16). catalytic subunit. Two contain sites for phosphorylation by PKA, which is activated by cyclic AMP. The remaining subunit is calmodulin, which binds Ca2+ when the cytosolic Ca2+ concentration rises. The regulatory subunits control the equilibrium between the active and inactive conformations of the catalytic subunit, with each phosphate and Ca2+ nudging the equilibrium toward the active conformation. How does this arrangement allow phosphorylase kinase to serve its role as an integrator protein for the multiple pathways that stimulate glycogen breakdown?


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Cell_Biology_Alberts. 15–17 The Wnt planar polarity signaling pathway normally ensures that each wing cell in Drosophila has a single hair. Overexpression of the Frizzled gene from a heat-shock promoter (hs-Fz) causes multiple hairs to grow from many cells (Figure Q15–5A). This phenotype is suppressed if hs-Fz is combined with a heterozygous deletion (DshΔ) of the Dishevelled gene (Figure Q15–5B). Do these results allow you to order the action of Frizzled and Dishevelled in the signaling pathway? If so, what is the order? Explain your reasoning. Figure Q15–5 Pattern of hair growth on wing cells in genetically different Drosophila (Problem 15–17). (From C.G. Winter et al., Cell 105:81–91, 2001. With permission from Elsevier.) Marks F, Klingmüller U & Müller-Decker K (2009) Cellular Signal Processing: An Introduction to the Molecular Mechanisms of Signal Transduction. New York: Garland Science. Lim W, Mayer B & Pawson T (2015) Cell Signaling: Principles and Mechanisms. New York: Garland Science.


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Cell_Biology_Alberts. Lim W, Mayer B & Pawson T (2015) Cell Signaling: Principles and Mechanisms. New York: Garland Science. Principles of Cell Signaling Alon U (2007) Network motifs: theory and experimental approaches. Nat. Rev. Genet. 8, 450–461. Ben-Shlomo I, Yu Hsu S, Rauch R et al (2003) Signaling receptome: a genomic and evolutionary perspective of plasma membrane receptors involved in signal transduction. Sci. STKE 187, RE9. Endicott JA, Noble ME & Johnson LN (2012) The structural basis for control of eukaryotic protein kinases. Annu. Rev. Biochem. 81, 587–613. Ferrell JE, Jr (2002) Self-perpetuating states in signal transduction: positive feedback, double-negative feedback and bistability. Curr. Opin. Cell Biol. 14, 140–148. Good MC, Zalatan JG & Lim WA (2011) Scaffold proteins: hubs for controlling the flow of cellular information. Science 332, 680–686. Ladbury JE & Arold ST (2012) Noise in cellular signaling pathways: causes and effects. Trends Biochem. Sci. 37, 173–178.


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Cell_Biology_Alberts. Ladbury JE & Arold ST (2012) Noise in cellular signaling pathways: causes and effects. Trends Biochem. Sci. 37, 173–178. Mehta S & Zhang J (2011) Reporting from the field: genetically encoded fluorescent reporters uncover signaling dynamics in living biological systems. Annu. Rev. Biochem. 80, 375–401. Pires-daSilva A & Sommer RJ (2003) The evolution of signalling pathways in animal development. Nature Rev. Genet. 4, 39–49. Rosse C, Linch M, Kermorgant S, Cameron AJ et al. (2010) PKC and the control of localized signal dynamics. Nat. Rev. Mol. Cell Biol. 11, 103–112. Scott JD & Pawson T (2009) Cell signaling in space and time: where proteins come together and when they’re apart. Science 326, 1220–1224. Seet BT, Dikic I, Zhou MM & Pawson T (2006) Reading protein modifications with interaction domains. Nat. Rev. Mol. Cell Biol. 7, 473–483.


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Cell_Biology_Alberts. Seet BT, Dikic I, Zhou MM & Pawson T (2006) Reading protein modifications with interaction domains. Nat. Rev. Mol. Cell Biol. 7, 473–483. Tyson JJ, Chen KC & Novak B (2003) Sniffers, buzzers, toggles and blinkers: dynamics of regulatory and signaling pathways in the cell. Curr. Opin. Cell Biol. 15, 221–231. Ubersax JA & Ferrell JE, Jr (2007) Mechanisms of specificity in protein phosphorylation. Nat. Rev. Mol. Cell Biol. 8, 530–541. Wittinghofer A & Vetter IR (2011) Structure-function relationships of the G domain, a canonical switch motif. Annu. Rev. Biochem. 80, 943–971. Audet M & Bouvier M (2012) Restructuring G-protein-coupled receptor activation. Cell 151, 14–23. Berridge MJ, Bootman MD & Roderick HL (2003) Calcium signalling: dynamics, homeostasis and remodelling. Nat. Rev. Mol. Cell Biol. 4, 517–529. Breer H (2003) Sense of smell: recognition and transduction of olfactory signals. Biochem. Soc. Trans. 31, 113–116.


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Cell_Biology_Alberts. Breer H (2003) Sense of smell: recognition and transduction of olfactory signals. Biochem. Soc. Trans. 31, 113–116. Hoeflich KP & Ikura M (2002) Calmodulin in action: diversity in target recognition and activation mechanisms. Cell 108, 739–742. Kamenetsky M, Middelhaufe S, Bank EM et al. (2006) Molecular details of cAMP generation in mammalian cells: a tale of two systems. J. Mol. Biol. 362, 623–639. McConnachie G, Langeberg LK & Scott JD (2006) AKAP signaling complexes: getting to the heart of the matter. Trends Mol. Med. 12, 317–323. Murad F (2006) Shattuck Lecture. Nitric oxide and cyclic GMP in cell signaling and drug development. N. Engl. J. Med. 355, 2003–2011. Parker PJ (2004) The ubiquitous phosphoinositides. Biochem. Soc. Trans. 32, 893–898. Rasmussen SG, DeVree BT, Zou Y et al. (2011) Crystal structure of the beta2 adrenergic receptor-Gs protein complex. Nature 477, 549–555. Rhee SG (2001) Regulation of phosphoinositide-specific phospholipase


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Cell_Biology_Alberts. Rhee SG (2001) Regulation of phosphoinositide-specific phospholipase C. Annu. Rev. Biochem. 70, 281–312. Robishaw JD & Berlot CH (2004) Translating G protein subunit diversity into functional specificity. Curr. Opin. Cell Biol. 16, 206–209. Rosenbaum DM, Rasmussen SG & Kobilka BK (2009) The structure and function of G-protein-coupled receptors. Nature 459, 356–363. Shenoy SK & Lefkowitz RJ (2011) beta-Arrestin-mediated receptor trafficking and signal transduction. Trends Pharmacol. Sci. 32, 521–533. Sorkin A & von Zastrow M (2009) Endocytosis and signalling: intertwining molecular networks. Nat. Rev. Mol. Cell Biol. 10, 609–622. Stratton MM, Chao LH, Schulman H & Kuriyan J (2013) Structural studies on the regulation of Ca2+/calmodulin dependent protein kinase II. Curr. Opin. Struct. Biol. 23, 292–301. Sung CH & Chuang JZ (2010) The cell biology of vision. J. Cell Biol. 190, 953–963. Willoughby D & Cooper DM (2008) Live-cell imaging of cAMP dynamics. Nat. Methods 5, 29–36.


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Cell_Biology_Alberts. Sung CH & Chuang JZ (2010) The cell biology of vision. J. Cell Biol. 190, 953–963. Willoughby D & Cooper DM (2008) Live-cell imaging of cAMP dynamics. Nat. Methods 5, 29–36. Jura N, Zhang X, Endres NF et al. (2011) Catalytic control in the EGF receptor and its connection to general kinase regulatory mechanisms. Mol. Cell 42, 9–22. Lemmon MA & Schlessinger J (2010) Cell signaling by receptor tyrosine kinases. Cell 141, 1117–1134. Massagué J (2012) TGFβ signalling in context. Nat. Rev. Mol. Cell Biol. 13, 616–630. Manning BD & Cantley LC (2007) AKT/PKB signaling: navigating downstream. Cell 129, 1261–1274. Mitin N, Rossman KL & Der CJ (2005) Signaling interplay in Ras superfamily function. Curr. Biol. 15, R563–R574. Pitulescu ME & Adams RH (2010) Eph/ephrin molecules—a hub for signaling and endocytosis. Genes Dev. 24, 2480–2492. Qi M & Elion EA (2005) MAP kinase pathways. J. Cell Sci. 118, 3569–3572.


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Cell_Biology_Alberts. Pitulescu ME & Adams RH (2010) Eph/ephrin molecules—a hub for signaling and endocytosis. Genes Dev. 24, 2480–2492. Qi M & Elion EA (2005) MAP kinase pathways. J. Cell Sci. 118, 3569–3572. Rawlings JS, Rosler KM & Harrison DA (2004) The JAK/STAT signaling pathway. J. Cell Sci. 117, 1281–1283. Roskoski R, Jr (2004) Src protein-tyrosine kinase structure and regulation. Biochem. Biophys. Res. Commun. 324, 1155–1164. Schwartz MA & Madhani HD (2004) Principles of MAP kinase signaling specificity in Saccharomyces cerevisiae. Annu. Rev. Genet. 38, 725–748. Shaw RJ & Cantley LC (2006) Ras, PI(3)K and mTOR signalling controls tumour cell growth. Nature 44, 424–430. Zoncu R, Efeyan A & Sabatini DM (2011) mTOR: from growth signal integration to cancer, diabetes and ageing. Nat. Rev. Mol. Cell Biol. 12, 21–35. Bray SJ (2006) Notch signalling: a simple pathway becomes complex. Nat. Rev. Mol. Cell Biol. 7, 678–689.


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Cell_Biology_Alberts. Bray SJ (2006) Notch signalling: a simple pathway becomes complex. Nat. Rev. Mol. Cell Biol. 7, 678–689. Briscoe J & Therond PP (2013) The mechanisms of Hedgehog signalling and its roles in development and disease. Nat. Rev. Mol. Cell Biol. 14, 416–429. Hoffmann A & Baltimore D (2006) Circuitry of nuclear factor κB signaling. Immunol. Rev. 210, 171–186. Huang P, Chandra V & Rastinejad F (2010) Structural overview of the nuclear receptor superfamily: insights into physiology and therapeutics. Annu. Rev. Physiol. 72, 247–272. Markson JS & O’Shea EK (2009) The molecular clockwork of a protein-based circadian oscillator. FEBS Lett. 583, 3938–3947. Mohawk JA, Green CB & Takahashi JS (2012) Central and peripheral circadian clocks in mammals. Annu. Rev. Neurosci. 35, 445–462. Niehrs C (2012) The complex world of WNT receptor signalling. Nat. Rev. Mol. Cell Biol. 13, 767–779.


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Cell_Biology_Alberts. Niehrs C (2012) The complex world of WNT receptor signalling. Nat. Rev. Mol. Cell Biol. 13, 767–779. Chen M, Chory J & Fankhauser C (2004) Light signal transduction in higher plants. Annu. Rev. Genet. 38, 87–117. Dievart A & Clark SE (2004) LRR-containing receptors regulating plant development and defense. Development 131, 251–261. Kim TW & Wang ZY (2010) Brassinosteroid signal transduction from receptor kinases to transcription factors. Annu. Rev. Plant Biol. 61, 681–704. Mockaitis K & Estelle M (2008) Auxin receptors and plant development: a new signaling paradigm. Annu. Rev. Cell Dev. Biol. 24, 55–80. Stepanova AN & Alonso JM (2009) Ethylene signaling and response: where different regulatory modules meet. Curr. Opin. Plant Biol. 12, 548–555. The Cytoskeleton 16


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Cell_Biology_Alberts. Stepanova AN & Alonso JM (2009) Ethylene signaling and response: where different regulatory modules meet. Curr. Opin. Plant Biol. 12, 548–555. The Cytoskeleton 16 For cells to function properly, they must organize themselves in space and interact mechanically with each other and with their environment. They have to be correctly shaped, physically robust, and properly structured internally. Many have to change their shape and move from place to place. All cells have to be able to rearrange their internal components as they grow, divide, and adapt to changing circumstances. These spatial and mechanical functions depend on a remarkable system of filaments called the cytoskeleton (Figure 16–1).


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Cell_Biology_Alberts. The cytoskeleton’s varied functions depend on the behavior of three families of protein filaments—actin filaments, microtubules, and intermediate filaments. Each type of filament has distinct mechanical properties, dynamics, and biological roles, but all share certain fundamental features. Just as we require our ligaments, bones, and muscles to work together, so all three cytoskeletal filament systems must normally function collectively to give a cell its strength, its shape, and its ability to move.


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Cell_Biology_Alberts. In this chapter, we describe the function and conservation of the three main filament systems. We explain the basic principles underlying filament assembly and disassembly, and how other proteins interact with the filaments to alter their dynamics, enabling the cell to establish and maintain internal order, to shape and remodel its surface, and to move organelles in a directed manner from one place to another. Finally, we discuss how the integration and regulation of the cytoskeleton allows a cell to move to new locations.


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Cell_Biology_Alberts. The three major cytoskeletal filaments are responsible for different aspects of the cell’s spatial organization and mechanical properties. Actin filaments determine the shape of the cell’s surface and are necessary for whole-cell locomotion; they also drive the pinching of one cell into two. Microtubules determine the positions of membrane-enclosed organelles, direct intracellular transport, and form the mitotic spindle that segregates chromosomes during cell division. Intermediate filaments provide mechanical strength. All of these cytoskeletal filaments interact with hundreds of accessory proteins that regulate and link the filaments to other cell components, as well as to each other. The accessory proteins are essential for the controlled assembly of the cytoskeletal filaments in particular locations, and they include the motor proteins, remarkable molecular machines that convert the energy of ATP hydrolysis into mechanical force that can either move organelles along the filaments


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Cell_Biology_Alberts. locations, and they include the motor proteins, remarkable molecular machines that convert the energy of ATP hydrolysis into mechanical force that can either move organelles along the filaments or move the filaments themselves.


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Cell_Biology_Alberts. In this section, we discuss the general features of the proteins that make up the filaments of the cytoskeleton. We focus on their ability to form intrinsically Figure 16–1 The cytoskeleton. (A) A cell in culture has been fixed and labeled to show its cytoplasmic arrays of microtubules (green) and actin filaments (red). (B) This dividing cell has been labeled to show its spindle microtubules (green) and surrounding cage of intermediate filaments (red). The DNA in both cells is labeled in blue. (A, courtesy of Albert Tousson; B, courtesy of Conly Rieder.) polarized and self-organized structures that are highly dynamic, allowing the cell to rapidly modify cytoskeletal structure and function under different conditions. Cytoskeletal Filaments Adapt to Form Dynamic or Stable Structures


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Cell_Biology_Alberts. Cytoskeletal Filaments Adapt to Form Dynamic or Stable Structures Cytoskeletal systems are dynamic and adaptable, organized more like ant trails than interstate highways. A single trail of ants may persist for many hours, extending from the ant nest to a delectable picnic site, but the individual ants within the trail are anything but static. If the ant scouts find a new and better source of food, or if the picnickers clean up and leave, the dynamic structure adapts with astonishing rapidity. In a similar way, large-scale cytoskeletal structures can change or persist, according to need, lasting for lengths of time ranging from less than a minute up to the cell’s lifetime. But the individual macromolecular components that make up these structures are in a constant state of flux. Thus, like the alteration of an ant trail, a structural rearrangement in a cell requires little extra energy when conditions change.


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Cell_Biology_Alberts. Regulation of the dynamic behavior and assembly of cytoskeletal filaments allows eukaryotic cells to build an enormous range of structures from the three basic filament systems. The micrographs in Panel 16–1 illustrate some of these structures. Actin filaments underlie the plasma membrane of animal cells, providing strength and shape to its thin lipid bilayer. They also form many types of cell-surface projections. Some of these are dynamic structures, such as the lamellipodia and filopodia that cells use to explore territory and move around. More stable arrays allow cells to brace themselves against an underlying substratum and enable muscle to contract. The regular bundles of stereocilia on the surface of hair cells in the inner ear contain stable bundles of actin filaments that tilt as rigid rods in response to sound, and similarly organized microvilli on the surface of intestinal epithelial cells vastly increase the apical cell-surface area to enhance nutrient absorption. In


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Cell_Biology_Alberts. as rigid rods in response to sound, and similarly organized microvilli on the surface of intestinal epithelial cells vastly increase the apical cell-surface area to enhance nutrient absorption. In plants, actin filaments drive the rapid streaming of cytoplasm inside cells.


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Cell_Biology_Alberts. Microtubules, which are frequently found in a cytoplasmic array that extends to the cell periphery, can quickly rearrange themselves to form a bipolar mitotic spindle during cell division. They can also form cilia, which function as motile whips or sensory devices on the surface of the cell, or tightly aligned bundles that serve as tracks for the transport of materials down long neuronal axons. In plant cells, organized arrays of microtubules help to direct the pattern of cell wall synthesis, and in many protozoans they form the framework upon which the entire cell is built. Intermediate filaments line the inner face of the nuclear envelope, forming a protective cage for the cell’s DNA; in the cytosol, they are twisted into strong cables that can hold epithelial cell sheets together or help nerve cells to extend long and robust axons, and they allow us to form tough appendages such as hair and fingernails.


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Cell_Biology_Alberts. An important and dramatic example of rapid reorganization of the cytoskeleton occurs during cell division, as shown in Figure 16–2 for a fibroblast growing in a tissue-culture dish. After the chromosomes have replicated, the interphase microtubule array that spreads throughout the cytoplasm is reconfigured into the bipolar mitotic spindle, which segregates the two copies of each chromosome into daughter nuclei. At the same time, the specialized actin structures that enable the fibroblast to crawl across the surface of the dish rearrange so that the cell stops moving and assumes a more spherical shape. Actin and its associated motor protein myosin then form a belt around the middle of the cell, the contractile ring, which constricts like a tiny muscle to pinch the cell in two. When division is complete, the cytoskeletons of the two daughter fibroblasts reassemble into their interphase structures to convert the two rounded-up daughter cells into smaller versions of the flattened,


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Cell_Biology_Alberts. is complete, the cytoskeletons of the two daughter fibroblasts reassemble into their interphase structures to convert the two rounded-up daughter cells into smaller versions of the flattened, crawling mother cell.


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Cell_Biology_Alberts. Many cells require rapid cytoskeletal rearrangements for their normal functioning during interphase as well. For example, the neutrophil, a type of white Actin flaments(also known as microflaments) are helical polymers of the protein actin. They are fexible structures with a diameter of 8 nm that organize into a variety of linear bundles, two-dimensional networks, and three-dimensional gels. Although actin flaments are dispersed throughout the cell, they are most highly concentrated in the cortex, just beneath the plasma membrane. (i) Single actin flament; (ii) microvilli; (iii) stress fbers (red) terminating in focal adhesions (green); (iv) striated muscle. Micrographs courtesy of R. Craig (i and iv); P.T. Matsudaira and D.R. Burgess (ii); K. Burridge (iii). Microtubulesare long, hollow cylinders made of the protein tubulin.


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Cell_Biology_Alberts. Micrographs courtesy of R. Craig (i and iv); P.T. Matsudaira and D.R. Burgess (ii); K. Burridge (iii). Microtubulesare long, hollow cylinders made of the protein tubulin. With an outer diameter of 25 nm, they are much more rigid than actin flaments. Microtubules are long and straight and frequently have one end attached to a microtubule-organizing center (MTOC) called a centrosome. (i) Single microtubule; (ii) cross section at the base of three (green) and organelles (red); (iv) ciliated protozoan. Micrographs courtesy of R. Wade (i); D.T. Woodrow and R.W. Linck (ii); D. Shima (iii); D. Burnette (iv).


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Cell_Biology_Alberts. Micrographs courtesy of R. Wade (i); D.T. Woodrow and R.W. Linck (ii); D. Shima (iii); D. Burnette (iv). Intermediate flaments are ropelike fbers with a diameter of about 10 nm; they are made of intermediate flament proteins, which constitute a large and heterogeneous family. One type of intermediate flament forms a meshwork called the nuclear lamina just beneath the inner nuclear membrane. Other types extend across the cytoplasm, giving cells mechanical strength. In an epithelial tissue, they span the cytoplasm from one cell–cell junction to another, thereby strengthening the entire epithelium. (i) Individual intermediate flaments; (ii) Intermediate flaments (blue) in neurons and (iii) epithelial cell; (iv) nuclear lamina. Micrographs courtesy of R. Quinlan (i); N. L. Kedersha (ii); M. Osborn (iii); U. Aebi (iv).


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Cell_Biology_Alberts. Micrographs courtesy of R. Quinlan (i); N. L. Kedersha (ii); M. Osborn (iii); U. Aebi (iv). blood cell, chases and engulfs bacterial and fungal cells that accidentally gain access to the normally sterile parts of the body, as through a cut in the skin. Like most crawling cells, neutrophils advance by extending a protrusive structure filled with newly polymerized actin filaments. When the elusive bacterial prey moves in a different direction, the neutrophil is poised to reorganize its polarized protrusive structures within seconds (Figure 16–3). The Cytoskeleton Determines Cellular Organization and Polarity


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Cell_Biology_Alberts. The Cytoskeleton Determines Cellular Organization and Polarity In cells that have achieved a stable, differentiated morphology—such as mature neurons or epithelial cells—the dynamic elements of the cytoskeleton must also provide stable, large-scale structures for cellular organization. On specialized epithelial cells that line organs such as the intestine and the lung, cytoskeletal-based cell-surface protrusions including microvilli and cilia are able to maintain a constant location, length, and diameter over the entire lifetime of the cell. For the actin bundles at the cores of microvilli on intestinal epithelial cells, this is only a few days. But the actin bundles at the cores of stereocilia on the hair cells of the inner ear must maintain their stable organization for the entire lifetime of the animal, since these cells do not turn over. Nonetheless, the individual actin filaments


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Cell_Biology_Alberts. Figure 16–2 Diagram of changes in cytoskeletal organization associated with cell division. The crawling fibroblast drawn here has a polarized, dynamic actin cytoskeleton (shown in red) that assembles lamellipodia and filopodia to push its leading edge toward the right. The polarization of the actin cytoskeleton is assisted by the microtubule cytoskeleton (green), consisting of long microtubules that emanate from a single microtubuleorganizing center located in front of the nucleus. When the cell divides, the polarized microtubule array rearranges to form a bipolar mitotic spindle, which is responsible for aligning and then segregating the duplicated chromosomes (brown). The actin filaments form a contractile ring at the center of the cell that pinches the cell in two after the chromosome segregation. After cell division is complete, the two daughter cells reorganize both the microtubule and actin cytoskeletons into smaller versions of those that were present in the mother cell,


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Cell_Biology_Alberts. segregation. After cell division is complete, the two daughter cells reorganize both the microtubule and actin cytoskeletons into smaller versions of those that were present in the mother cell, enabling them to crawl their separate ways.


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Cell_Biology_Alberts. Figure 16–3 A neutrophil in pursuit of bacteria. In this preparation of human blood, a clump of bacteria (white arrow) is about to be captured by a neutrophil. As the bacteria move, the neutrophil quickly reassembles the dense actin network at its leading edge (highlighted in red) to push toward the location of the bacteria (Movie 16.1). Rapid disassembly and reassembly of the actin cytoskeleton in this cell enables it to change its orientation and direction of movement within a few minutes. (From a video recorded by David Rogers.) remain strikingly dynamic and are continuously remodeled and replaced every 48 hours, even within stable cell-surface structures that persist for decades.


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Cell_Biology_Alberts. Besides forming stable, specialized cell-surface protrusions, the cytoskeleton is also responsible for large-scale cellular polarity, enabling cells to tell the difference between top and bottom, or front and back. The large-scale polarity information conveyed by cytoskeletal organization is often maintained over the lifetime of the cell. Polarized epithelial cells use organized arrays of microtubules, actin filaments, and intermediate filaments to maintain the critical differences between the apical surface and the basolateral surface. They also must maintain strong adhesive contacts with one another to enable this single layer of cells to serve as an effective physical barrier (Figure 16–4).


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Cell_Biology_Alberts. Cytoskeletal filaments can reach from one end of the cell to the other, spanning tens or even hundreds of micrometers. Yet the individual protein molecules that form the filaments are only a few nanometers in size. The cell builds the filaments by assembling large numbers of the small subunits, like building a skyscraper out of bricks. Because these subunits are small, they can diffuse rapidly in the cytosol, whereas the assembled filaments cannot. In this way, cells can undergo rapid structural reorganizations, disassembling filaments at one site and reassembling them at another site far away.


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Cell_Biology_Alberts. Actin filaments and microtubules are built from subunits that are compact and globular—actin subunits for actin filaments and tubulin subunits for microtubules—whereas intermediate filaments are made up of smaller subunits that are themselves elongated and fibrous. All three major types of cytoskeletal filaments form as helical assemblies of subunits (see Figure 3–22) that self-associate, using a combination of end-to-end and side-to-side protein contacts. Differences in the structures of the subunits and the strengths of the attractive forces between microvillus terminal web of actin Figure 16–4 Organization of the cytoskeleton in polarized epithelial cells.


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Cell_Biology_Alberts. All the components of the cytoskeleton cooperate to produce the characteristic shapes of specialized cells, including the epithelial cells that line the small intestine, diagrammed here. At the apical (upper) surface, facing the intestinal lumen, bundled actin filaments (red) form microvilli that increase the cell surface area available for absorbing nutrients from food. Below the microvilli, a circumferential band of actin filaments is connected to cell–cell adherens junctions that anchor the cells to each other. Intermediate filaments (blue) are anchored to other kinds of adhesive structures, including desmosomes and hemidesmosomes, that connect the epithelial cells into a sturdy sheet and attach them to the underlying extracellular matrix; these structures are discussed in Chapter 19. Microtubules (green) run vertically from the top of the cell to the bottom and provide a global coordinate system that enables the cell to direct newly synthesized components to their proper


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Cell_Biology_Alberts. 19. Microtubules (green) run vertically from the top of the cell to the bottom and provide a global coordinate system that enables the cell to direct newly synthesized components to their proper locations.


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Cell_Biology_Alberts. them produce important differences in the stability and mechanical properties of each type of filament. Whereas covalent linkages between their subunits hold together the backbones of many biological polymers—including DNA, RNA, and proteins—it is weak noncovalent interactions that hold together the three types of cytoskeletal polymers. Consequently, their assembly and disassembly can occur rapidly, without covalent bonds being formed or broken.


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Cell_Biology_Alberts. The subunits of actin filaments and microtubules are asymmetrical and bind to one another head-to-tail so that they all point in one direction. This subunit polarity gives the filaments structural polarity along their length, and makes the two ends of each polymer behave differently. In addition, actin and tubulin subunits are both enzymes that catalyze the hydrolysis of a nucleoside triphosphate— ATP and GTP, respectively. As we discuss later, the energy derived from nucleotide hydrolysis enables the filaments to remodel rapidly. By controlling when and where actin and microtubules assemble, the cell harnesses the polar and dynamic properties of these filaments to generate force in a specific direction, to move the leading edge of a migrating cell forward, for example, or to pull chromosomes apart during cell division. In contrast, the subunits of intermediate filaments are symmetrical, and thus do not form polarized filaments with two different ends. Intermediate filament subunits


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Cell_Biology_Alberts. apart during cell division. In contrast, the subunits of intermediate filaments are symmetrical, and thus do not form polarized filaments with two different ends. Intermediate filament subunits also do not catalyze the hydrolysis of nucleotides. Nevertheless, intermediate filaments can be disassembled rapidly when required. In mitosis , for example, kinases phosphorylate the subunits, leading to their dissociation.


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Cell_Biology_Alberts. Cytoskeletal filaments in living cells are not built by simply stringing subunits together in single file. A thousand tubulin subunits lined up end-to-end, for example, would span the diameter of a small eukaryotic cell, but a filament formed in this way would lack the strength to avoid breakage by ambient thermal energy, unless each subunit in the filament was bound extremely tightly to its two neighbors. Such tight binding would limit the rate at which the filaments could disassemble, making the cytoskeleton a static and less useful structure. To provide both strength and adaptability, microtubules are built of 13 protofilaments—linear strings of subunits joined end-to-end—that associate with one another laterally to form a hollow cylinder. The addition or loss of a subunit at the end of one protofilament makes or breaks a small number of bonds. In contrast, loss of a subunit from the middle of the filament requires breaking many more bonds, while breaking it in two requires


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Cell_Biology_Alberts. end of one protofilament makes or breaks a small number of bonds. In contrast, loss of a subunit from the middle of the filament requires breaking many more bonds, while breaking it in two requires breaking bonds in multiple protofilaments all at the same time (Figure 16–5). The greater energy required to break multiple noncovalent bonds simultaneously allows microtubules to resist thermal breakage, while allowing rapid subunit addition and loss at the filament ends. Helical actin filaments are much thinner and therefore require much less energy to break. However, multiple actin filaments are often bundled together inside cells, providing mechanical strength, while allowing dynamic behavior of filament ends.


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Cell_Biology_Alberts. As with other specific protein–protein interactions, many hydrophobic interactions and noncovalent bonds hold the subunits in a cytoskeletal filament together (see Figure 3–4). The locations and types of subunit–subunit contacts differ for the different filaments. Intermediate filaments, for example, assemble by forming strong lateral contacts between α-helical coiled-coils, which extend over most of the length of each elongated fibrous subunit. Because the individual subunits are staggered in the filament, intermediate filaments form strong, ropelike structures that tolerate stretching and bending to a greater extent than do either actin filaments or microtubules (Figure 16–6).


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Cell_Biology_Alberts. The cell regulates the length and stability of its cytoskeletal filaments, as well as their number and geometry. It does so largely by regulating their attachments to one another and to other components of the cell, so that the filaments can form a wide variety of higher-order structures. Direct covalent modification of the filament subunits regulates some filament properties, but most of the regulation is performed by hundreds of accessory proteins that determine the spatial SINGLE PROTOFILAMENT: THERMALLY UNSTABLE


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Cell_Biology_Alberts. MULTIPLE PROTOFILAMENTS: THERMALLY STABLE distribution and the dynamic behavior of the filaments, converting information received through signaling pathways into cytoskeletal action. These accessory proteins bind to the filaments or their subunits to determine the sites of assembly of new filaments, to regulate the partitioning of polymer proteins between filament and subunit forms, to change the kinetics of filament assembly and disassembly, to harness energy to generate force, and to link filaments to one another or to other cell structures such as organelles and the plasma membrane. In these processes, the accessory proteins bring cytoskeletal structure under the control of extracellular and intracellular signals, including those that trigger the dramatic transformations of the cytoskeleton that occur during each cell cycle. Acting together, the accessory proteins enable a eukaryotic cell to maintain a highly organized but flexible internal structure and, in many cases, to move.


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Cell_Biology_Alberts. Figure 16–5 The thermal stability of cytoskeletal filaments with dynamic ends. A protofilament consisting of a single strand of subunits is thermally unstable, since breakage of a single bond between subunits is sufficient to break the filament. In contrast, formation of a cytoskeletal filament from more than one protofilament allows the ends to be dynamic, while enabling the filaments themselves to be resistant to thermal breakage. In a microtubule, for example, removing a single subunit dimer from the end of the filament requires breaking noncovalent bonds with a maximum of three other subunits, whereas fracturing the filament in the middle requires breaking noncovalent bonds in all thirteen protofilaments.


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Cell_Biology_Alberts. Figure 16–6 Flexibility and stretch in an intermediate filament. Intermediate filaments are formed from elongated fibrous subunits with strong lateral contacts, resulting in resistance to stretching forces. When a tiny mechanical probe is dragged across an intermediate filament, the filament is stretched over three times its length before it breaks, as illustrated by the fluorescently labeled filaments in the photomicrographs. This technique is termed atomic force microscopy (see Figure 9–33). (Adapted from L. Kreplak et al., J. Mol. Biol. 354:569–577, 2005. With permission from Elsevier.)


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Cell_Biology_Alberts. Among the most fascinating proteins that associate with the cytoskeleton are the motor proteins. These proteins bind to a polarized cytoskeletal filament and use the energy derived from repeated cycles of ATP hydrolysis to move along it. Dozens of different motor proteins coexist in every eukaryotic cell. They differ in the type of filament they bind to (either actin or microtubules), the direction in which they move along the filament, and the “cargo” they carry. Many motor proteins carry membrane-enclosed organelles—such as mitochondria, Golgi stacks, or secretory vesicles—to their appropriate locations in the cell. Other motor proteins cause cytoskeletal filaments to exert tension or to slide against each other, generating the force that drives such phenomena as muscle contraction, ciliary beating, and cell division.


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Cell_Biology_Alberts. Cytoskeletal motor proteins that move unidirectionally along an oriented polymer track are reminiscent of some other proteins and protein complexes discussed elsewhere in this book, such as DNA and RNA polymerases, helicases, and ribosomes. All of these proteins have the ability to use chemical energy to propel themselves along a linear track, with the direction of sliding dependent on the structural polarity of the track. All of them generate motion by coupling nucleoside triphosphate hydrolysis to a large-scale conformational change (see Figure 3–75). Bacterial Cell Organization and Division Depend on Homologs of Eukaryotic Cytoskeletal Proteins


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Cell_Biology_Alberts. Bacterial Cell Organization and Division Depend on Homologs of Eukaryotic Cytoskeletal Proteins While eukaryotic cells are typically large and morphologically complex, bacterial cells are usually only a few micrometers long and assume simple shapes such as spheres or rods. Bacteria also lack elaborate networks of intracellular membrane-enclosed organelles. Historically, biologists assumed that a cytoskeleton was not necessary in such simple cells. We now know, however, that bacteria contain homologs of all three of the eukaryotic cytoskeletal filaments. Furthermore, bacterial actins and tubulins are more diverse than their eukaryotic versions, both in the types of assemblies they form and in the functions they carry out.


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Cell_Biology_Alberts. Nearly all bacteria and many archaea contain a homolog of tubulin called FtsZ, which can polymerize into filaments and assemble into a ring (called the Z-ring) at the site where the septum forms during cell division (Figure 16–7). Although the Z-ring persists for many minutes, the individual filaments within it are highly dynamic, with an average filament half-life of about thirty seconds. As the bacte rium divides, the Z-ring becomes smaller until it has completely disassembled. FtsZ filaments in the Z-ring are thought to generate a bending force that drives the membrane invagination necessary to complete cell division. The Z-ring may also serve as a site for localization of enzymes required for building the septum between the two daughter cells.


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Cell_Biology_Alberts. Many bacteria also contain homologs of actin. Two of these, MreB and Mbl, are found primarily in rod-shaped or spiral-shaped cells where they assemble to form dynamic patches that move circumferentially along the length of the cell (Figure 16–8A). These proteins contribute to cell shape by serving as a scaffold to direct the synthesis of the peptidoglycan cell wall, in much the same way that microtubules help organize the synthesis of the cellulose cell wall in higher


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Cell_Biology_Alberts. Figure 16–7 The bacterial FtsZ protein, a tubulin homolog in prokaryotes. (A) A band of FtsZ protein forms a ring in a dividing bacterial cell. This ring has been labeled by fusing the FtsZ protein to green fluorescent protein (GFP), which allows it to be observed in living E. coli cells with a fluorescence microscope. (B) FtsZ filaments and circles, formed in vitro, as visualized using electron microscopy. (C) Dividing chloroplasts (red) from a red alga also cleave using a protein ring made from FtsZ (yellow). (A, from X. Ma, D.W. Ehrhardt and W. Margolin, Proc. Natl Acad. Sci. USA 93:12998–13003, 1996; B, from H.P. Erickson et al., Proc. Natl Acad. Sci. USA 93:519–523, 1996. Both with permission from National Academy of Sciences; C, from S. Miyagishima et al., Plant Cell 13:2257–2268, 2001, with permission from (C)American Society of Plant Biologists.) 1 µm


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Cell_Biology_Alberts. S. Miyagishima et al., Plant Cell 13:2257–2268, 2001, with permission from (C)American Society of Plant Biologists.) 1 µm Figure 16–8 Actin homologs in bacteria determine cell shape. (A) The MreB protein forms abundant patches made up of many short, interwoven linear or helical filaments that are seen to move circumferentially along the length of the bacterium and are associated with sites of cell wall synthesis. (B) The common soil bacterium Bacillus subtilis normally forms cells with a regular rodlike shape when viewed by scanning electron microscopy (left). In contrast, B. subtilis cells lacking the actin homolog MreB or Mbl grow in distorted or twisted shapes and eventually die (center and right). (A, from P. Vats and L. Rothfield, Proc. Natl Acad. Sci. USA 104:17795–17800, 2007. With permission from National Academy of Sciences; B, from A. Chastanet and


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Cell_Biology_Alberts. R. Carballido-Lopez, Front. Biosci. 4S:1582–1606, 2012. With permission Frontiers in Bioscience.) plant cells (see Figure 19–65). As with FtsZ, MreB and Mbl filaments are highly dynamic, with half-lives of a few minutes, and nucleotide hydrolysis accompanies the polymerization process. Mutations disrupting MreB or Mbl expression cause extreme abnormalities in cell shape and defects in chromosome segregation (Figure 16–8B).


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Cell_Biology_Alberts. Relatives of MreB and Mbl have more specialized roles. A particularly intriguing bacterial actin homolog is ParM, which is encoded by a gene on certain bacterial plasmids that also carry genes responsible for antibiotic resistance and cause the spread of multidrug resistance in epidemics. Bacterial plasmids typically encode all the gene products that are necessary for their own segregation, presumably as a strategy to ensure their inheritance and propagation in bacterial hosts following plasmid replication. ParM assembles into filaments that associate at each end with a copy of the plasmid, and growth of the ParM filament pushes the replicated plasmid copies apart (Figure 16–9). This spindle-like structure apparently arises from the selective stabilization of filaments that bind to specialized proteins recruited to the origins of replication on the plasmids. A distant relative of both tubulin and FtsZ, called TubZ, has a similar function in other bacterial species.


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Cell_Biology_Alberts. Thus, self-association of nucleotide-binding proteins into dynamic filaments is used in all cells, and the actin and tubulin families are very ancient, predating the split between the eukaryotic and bacterial kingdoms. At least one bacterial species, Caulobacter crescentus, appears to harbor a protein with significant structural similarity to the third major class of cytoskeletal filaments found in animal cells, the intermediate filaments. A protein called crescentin forms a filamentous structure that influences the unusual crescent shape of this species; when the gene encoding crescentin is deleted, the Caulobacter cells grow as straight rods (Figure 16–10). ParM plasmid origin of ParM ParR monomers replication flaments proteins Figure 16–9 Role of the actin homolog ParM in plasmid segregation in bacteria.


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Cell_Biology_Alberts. Figure 16–9 Role of the actin homolog ParM in plasmid segregation in bacteria. (A) Some bacterial drug-resistance plasmids (orange) encode an actin homolog, ParM, that will spontaneously nucleate to form small, dynamic filaments (green) throughout the bacterial cytoplasm. A second plasmid-encoded protein called ParR (blue) binds to specific DNA sequences in the plasmid and also stabilizes the dynamic ends of the ParM filaments. When the plasmid duplicates, both ends of the ParM filaments become stabilized, and the growing ParM filaments push the duplicated plasmids to opposite ends of the cell. (B) In these bacterial cells harboring a drug-resistance plasmid, the plasmids are labeled in red and the ParM protein in green. Left, a short ParM filament bundle connects the two daughter plasmids shortly after their duplication. Right, the fully assembled ParM filament has pushed the duplicated plasmids to the cell poles. (A, adapted from E.C. Garner,


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Cell_Biology_Alberts. C.S. Campbell and R.D. Mullins, Science 306:1021–1025, 2004; B, from J. Møller-Jensen et al., Mol. Cell 12:1477–1487, 2003. With permission from Elsevier.)


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Cell_Biology_Alberts. The cytoplasm of eukaryotic cells is spatially organized by a network of protein filaments known as the cytoskeleton. This network contains three principal types of filaments: actin filaments, microtubules, and intermediate filaments. All three types of filaments form as helical assemblies of subunits that self-associate using a combination of end-to-end and side-to-side protein contacts. Differences in the structure of the subunits and the manner of their self-assembly give the filaments different mechanical properties. Subunit assembly and disassembly constantly remodel all three types of cytoskeletal filaments. Actin and tubulin (the subunits of actin filaments and microtubules, respectively) bind and hydrolyze nucleoside triphosphates (ATP and GTP, respectively), and assemble head-to-tail to generate polarized filaments capable of generating force. In living cells, accessory proteins modulate the dynamics and organization of cytoskeletal filaments, resulting in complex events such


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Cell_Biology_Alberts. to generate polarized filaments capable of generating force. In living cells, accessory proteins modulate the dynamics and organization of cytoskeletal filaments, resulting in complex events such as cell division or migration, and generating elaborate cellular architecture to form polarized tissues such as epithelia. Bacterial cells also contain homologs of actin, tubulin, and intermediate filaments that form dynamic structures that help control cell shape and division.


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Cell_Biology_Alberts. The actin cytoskeleton performs a wide range of functions in diverse cell types. Each actin subunit, sometimes called globular or G-actin, is a 375-amino-acid polypeptide carrying a tightly associated molecule of ATP or ADP (Figure 16–11A). Actin is extraordinarily well conserved among eukaryotes. The amino acid sequences of actins from different eukaryotic species are usually about 90% identical. Small variations in actin amino acid sequence can cause significant functional differences: In vertebrates, for example, there are three isoforms of actin, termed α, β, and γ, that differ slightly in their amino acid sequences and have distinct functions. α-Actin is expressed only in muscle cells, while βand γ-actins are found together in almost all non-muscle cells. Actin Subunits Assemble Head-to-Tail to Create Flexible, Polar Filaments


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Cell_Biology_Alberts. Actin Subunits Assemble Head-to-Tail to Create Flexible, Polar Filaments Actin subunits assemble head-to-tail to form a tight, right-handed helix, forming a structure about 8 nm wide called filamentous or F-actin (Figure 16–11B and C). Because the asymmetrical actin subunits of a filament all point in the same direction, filaments are polar and have structurally different ends: a slower-growing minus end and a faster-growing plus end. The minus end is also referred to as the “pointed end” and the plus end as the “barbed end,” because of the “arrowhead” appearance of the complex formed between actin filaments and the motor protein myosin (Figure 16–12). Within the filament, the subunits are positioned with their nucleotide-binding cleft directed toward the minus end.


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Cell_Biology_Alberts. Individual actin filaments are quite flexible. The stiffness of a filament can be characterized by its persistence length, the minimum filament length at which random thermal fluctuations are likely to cause it to bend. The persistence length of an actin filament is only a few tens of micrometers. In a living cell, however, Figure 16–10 Caulobacter and crescentin. The sickle-shaped bacterium Caulobacter crescentus expresses a protein, crescentin, with a series of coiled-coil domains similar in size and organization to the domains of eukaryotic intermediate filaments. (A) The crescentin protein forms a fiber (labeled in red) that runs down the inner side of the curving bacterial cell wall. (B) When the gene is disrupted, the bacteria grow as straight rods (bottom). (From N. Ausmees, J.R. Kuhn and