Hello and welcome to the Grand Line Review, your source for everything One Piece. Recently some mildly interesting information has popped up in the form of a partially leaked article from V-Jump, which is a monthly publication that focuses on video game-based manga. But for whatever reason, in this edition we have a section on the one true manga, One Piece. And before I launch into it, I first need to give all credit to the Library of Ohara, who translated the leaked portion of the article, and there will be a link to him in the description below. But it's some pretty big stuff here, as it deals with the bounties of the Yonko. First up, the article poses a lovely little question, which is, what are the bounties of the Yonko? Hmm, intriguing. Immediately going on to state that the two monstrous individuals known as Big Mom and Kaito are priced at over two billion. Now we should take everything here with a healthy dose of skepticism, because this was not written by Oda himself. But it would have had to have been approved by him, so with that in mind, it seems to be quite implied that Big Mom and Kaito have bounties of less than three billion berries. And that would very much place them within Blackbeard range, which makes some sense I guess. But I need to stress that we cannot take any fact from such an implication. It is entirely possible that both of them have bounties of over three billion, but the only thing we know for sure is that they are over two billion. But next we have what is by far the most interesting portion to me concerning a certain shanks. And I'll read the translation word for word here, because it is of great semantic importance. Despite being the only Yonko not to have a devil fruit, he's powerful enough to easily fight against marine admiral level. His bounty is the number one highest. So despite a little fumble with the English language there, this is a pretty huge bombshell. First up in one fell swoop of a random article, out goes every theory in existence on shanks having a devil fruit. Which is a shame because there were some pretty cool ideas out there, but here we are.  I think that while I very much did enjoy the mystery of shanks and whether or not he was a fruit user being up in the air, it's nice to know for a fact that he's just a complete beast in his own right. The justification for his strength in this article is that he is able to fight against a marine admiral. And to that I say that's a pretty terrible measure of strength. I mean admiral are terrifyingly strong beings, don't get me wrong. But not even three of them plus the rest of the entirety of the marines were able to take down a single Yonko. So you'd figure that another Yonko should be able to take on an admiral. But just on this topic, the library of Ohara himself spoke on this matter telling us to keep in mind that what's written here is very vague, offering that the translation could more pragmatically be masterfully matched or something along those lines. I don't know though it still feels a lot like saying that Luffy is powerful enough to masterfully match Don Krieg. You know it's just a weird thing to say. Although I guess when you think about it they don't have a lot of options to compare shanks to. I guess they could have said he was powerful enough to masterfully match another Yonko. But that's pretty redundant given his position as a Yonko. So hey admirals why not? Actually you know what it might even be a direct reference to when shanks confronted Sakazuki at Marineford. And as for the final portion where it states that shanks bounty is the number one highest, well now that's pretty massive isn't it? Given that we know so little about shanks I've never really tried to put him on a bounty or power scale in comparison to that of the other Yonko. But prior to today if someone had put a gun to my head and told me to rank the Yonko by bounty value I probably would have put shanks below both Whitebeard and Kaido. But this little revelation has given me renewed interest in older red hair. You must be quite something to be considered more dangerous than these absolute monsters we've been examining recently in the series. But hey I suppose it's entirely possible that he knows where Raftel is so there's that. That's uh pretty important and stuff. But before people jump to conclusions regarding the wording of shanks's bounty being the number one highest let's take a moment to reflect on how incredibly vague this wording is. Yes it could mean that shanks has the highest bounty in the entire one piece world which would conflict with the previously conceived notion that that title would belong to Dragon being the most wanted man in the world. I mean I guess the most wanted may not necessarily mean the highest priced but I'm more inclined to believe that through the context of the article it is implying that shanks's bounty is number one amongst the Yonko rather than the highest in the world at large. Interestingly enough Luffy also gets a little feature of his own in this article and while he is called the newest emperor of the sea a question mark is strategically placed at the end of that statement implying that as we suspected his position isn't exactly confirmed yet. Some people consider him an emperor others don't like Blackbeard. Speaking of he also does appear in the article but his segment isn't on this leak. So there is entirely possibly a ton of information to come out when the publication is officially released and if anything is relevant then I'll be sure to keep you all informed. But if you enjoy this video and the content this channel produces in general then please do consider donating to the Grand Line review Patreon because the support of all of your amazing people is what continues to make this channel possible. Also do check out my Teespring store if you're interested in shirts hoodies and other miscellaneous items but the proceeds going to directly to support the channel as well and if you'd like to join the fun then please do head over to my discord server where a wide array of shenanigan retakes place on a daily basis. And finally please do comment with your thoughts on the Yonko bounty news. This has been the Grand Line review and I'll see you next time.


Kia ora, my name is Bisma and welcome to this week's Year Away Weekly episode. Today we're coming to you from the Grafton campus. Wookiee Fire Holder Week is all about student wellbeing and mental health for the students at Year Away. This week the university is hosting many relaxing and informative events. This includes yoga, paint sessions, free food, free access to the rec center and so much more. As part of the professional development program the university and women in engineering will be hosting a moldy cultural competency breakfast. Snacks and drinks will be provided to come along this Thursday 18th of May at 8 a.m to the Terraces Engineering Building. Join us for a free glow flow yoga session and move your body in a fun and non-judgmental environment. Come dressed in white or neon for the best glow effect and don't forget to bring your towel and water bottle. Join the Volunteer Club this Saturday 20th of May from 9 30 a.m to 11 30 a.m at the William Gamble Reserve for their planting day. Spend the morning planting and weeding and enjoy a delicious saucer schedule right at the end. Join us at the 2023 Interfaculty Basketball and Futsal Championships Finals. This will be on Friday 26th of May from 5 p.m. There will be free food and alongside crowd entertainment and activities. Come along and support your favorite faculty. Calling all nursing students, check out and sign up to UOA NASA. The club is all about supporting and celebrating nursing students. The Business School ASB Career Center is open Monday 1 p.m to 2 p.m for students to come in and drop their CVs, cover letters, linkedins and application forms. If you can't make the on-campus time slot there is the zoom option available. That's all from us this week. For more information and to contribute to your weekly, go to the what's on page at the University's website. Ka kite ano!


So, good afternoon everybody. It's my great pleasure to introduce the first distinguished visiting scientists program talk. And it's been a great honor to host Professor Christian Rosamund in my lab. And he is the speaker today. Christian is a really well known figure or person, a scientist studying SNAPTIC transmission throughout his career. And let me introduce his biography. So he is from Germany, and he originally studied pharmacology from Goethe University in Frankfurt, Germany. Then after that, a little bit unusual, I think at that time, he went over to the US to do his PhD at the Wollum Institute with Gary Westbrook. And after that, that's when I got to know him. He went to do a postdoc at the Salk Institute with Chuck Stevens. Yeah, this is way back in 1993, we're just chatting almost 30 years ago. And then subsequently he became a Helmholtz fellow and moved to Germany in Göttingen at the Max Planck Institute for biophysical chemistry with Dr. Alan Marti. And then he subsequently still staying at the MPI by physical chemistry. He became a principal investigator of Heisenberg fellow. And then in 2008, he moved to Baylor back to US now this time in Texas to take up an assistant professorship in the Department of Molecular and Human Genetics, and also in Department of Neuroscience. And then working through the ranks, he became a professor in 2008 at Baylor. And then soon after he moved back to Germany this time to Berlin at Charité, where he is currently to take up a professorship and being involved in multiple responsibilities at Charité. And he's currently a professor in the Center for Basic Sciences at the Institute of Neurophysiology. So apart from these very prestigious fellowships through which he's gained various positions. He's been awarded many grants, prestigious grants, and also since 2014, he's a member of a Board of Trustees of Schramm Foundation in Essen, Germany. And since 2015, he's serving as a scientific advisory board member in the Department of Biomedicine and University of Basel. And for in recognition of his outstanding scientific contributions in 2019, he was elected as a member of National Academy of Sciences Leopoldina in Germany, which is the highest scientific distinction bestowed by a German institution. And he's, as I mentioned, he's an expert in synaptic transmission, and he's really well known for his elegant work throughout his career. I think my very first strong impression was this really highly cited 1993 science paper that he produced as a PhD student, taking advantage of his pharmacology background and really having an elegant method to estimate the diversity of presnaptic strengths, which is a method that's been used throughout, I won't go into the details, but you will hear more from Christian. And I think my favorite is the his identification of cytoskeletal modulation of ion channel. And since then he's been involved in regulation of neurotransmitter release that's very crucial for accuracy of information transmission. And so without further ado, Christian, please. Well, thank you Yuki for introducing me so so graciously. So many honors I nice to hear about. Yeah, I just don't. Yeah, we love the signs. I think this is always the great part. And, you know, I want to maybe start before I go into my research topic would really like to thank OIST to allow me to do my sabbatical here since last September in Yuki's lab. And this was done through the distinguished distinguished visiting scholar program. So again, thank you so much for making that possible for me to come here. I particularly would like to thank Jonas Fischer and his team to to help me go through the moving here part which is, you know, it's it's it was quite a little interesting adventure not not complicated but very interesting and it still remains to be. And yeah, it's a very great honor to be in this very stimulating environment this various prestigious Institute. And yeah, I'm still looking forward for three more months of of interaction with you guys so. Yeah, so what I want to talk to you about is is really kind of at the core of what my laboratories interested in is how synapses actually communicate what's how do they actually do their job. And the, let me just maybe start a bit more broader. The brain, you know, we all know this is probably the most complicated structure in the universe. We know when we look at the anatomy of the human brain. There are approximately 1000 different sub areas within the brain, executing various functions that we performing every day when we think when we sleep when we eat when we have feelings, etc. And that of course is mediated by a whole large host of cells that underlie these structures. There are, of course, mainly to mention the neurons that are astonishingly 86 billion neurons in the human brain. Interestingly, also we see that the number of glia cells and this is something that brought me here to to to for my visit make make almost the same up the same number of cells in the brain and studying synaptic transmission. We really have to start thinking about not start thinking I have to start thinking about it. Yuki is doing that for many years already. What the glia is doing and the interaction and helping synapses to actually do what they're supposed to do. Now, this is just some statistics of the human near cortex. You, when you just look at a microliter of brain volume, you find in there not only about 40,000 neurons, but each of these neurons make approximately 15,000 synaptic connections to other neurons. At the same time, received a similar number of synapses are making this an incredible complex network of interactions. And we find in this microliter of brain volume, four kilometers of axon lengths, which just tells you the complexity of interactions. And if you are just looking at like a cube, that is about 2000 times smaller of this one microliter which is 0.5 nano liter of volume. And you make a reconstruction of all the neurons and connections in the brain in this in this volume of cortex. You can appreciate the complexity of this structures. So you see here in blue axons, later in cyan, you will see the synapses. We're just sort of diving into this tissue through a vessel. See the synapses colored in cyan. And it's just amazing to think about 80 by 80 by 80 micron sized cube how complex that structures here you start seeing an axon. And a dendrite and the synaptic connections in between. This is an individual synapse and of course you can see they are in this cube about 80,000 of those connections. And so as you can, you can think in a way that the brain is a humongous, humongous computational organ that enables us to communicate information across neurons across synapses. So my interest has been, you know, thinking of the brain. Of course, you have to think about the areas and neurons, how they're connected, how the neurons are electrically active. And I have been particularly interested in the synapses, how they actually convey that information from the pre synaptic side to the post synaptic side via action potential and use neuro transmitter release that in turn then activates post synaptic receptors. So just a kind of a scheme of this. One of the home walks of synaptic transmission is really it's incredible speed. But let me show you first what happens really at the synapse we have a pre synaptic depolarization induced by the incoming action potential. You have council channels sitting right next to it that open council fluxes in and then you have fusion competent vesicles that are sort of sitting right next to the membrane, waiting for the, the, the, the go signal. This relates to the stop and go of my title, they're waiting for the go signal to then actually fuses the plasma membrane releasing its neuro transmitter content into the synaptic cleft, and then activating the post synaptic receptors. To open it's, it's gate. So cutting on and cut irons in case of photomate receptors, sort of flow into the post synaptic dendrite, we selecting the pre synaptic electrical activity as a post synaptic depolarization. So here's just to show you the speed questions of when you when you look at functional of neurons like to physiology is a really important technique to do this. You see here the pre synaptic action potential recorded from a from a nerve terminal, and then you see subsequently in the post synaptic membrane, either by looking at current clump recordings of depolarization of the membrane. Or when you look at voltage clump recordings in inward current that occurs on really in a millisecond time scale. So if you then blow this up in terms of the time scale you see here the pre synaptic action potential, you see the pre synaptic counts in current and then you see the post synaptic response. You can start appreciating the speed between the pre synaptic and the post synaptic signal occurring within a millisecond or so depends a little bit on the synapse you're looking at. But this is very important because neurons fire in the brain at very very high frequencies, and the precision of this firing of the neuron is an extremely valuable information content. And so synapses job is to maintain that pre temporal precision, and therefore synapses have to be very very fast. And so that's one of the things that fascinated me in this area. How can this cell biological process of vesicle fusion induced by calcium influx take place in a microsecond time scale. Here perhaps the probably the most fast the fastest cell biological process that I know of but you know given this diverse auditorium yet always maybe someone comes up with something faster. I'm very curious to hear about it but anyway it's one of the fastest things that happen in biology. But the other thing that you have to keep in mind with synaptic transmission is not only speed, but you also have to keep in mind that synapses are have have a very, very precious content that are easily be consumed each other synaptic vesicles. The pre synaptic terminal and the nervous central nervous system has only a few vesicles at hand fusion ready ready vesicles that can be easily consumed if you think of a fifth or sixth action potential. And then once those vesicles are gone they need to be refilled. And so really as an issue of supply demand that the nerves the pre synaptic nervous pre synaptic nerve terminal has to face. When you look at ongoing firing of synaptic transmission, you see that the amplitude of the response for example can go into their knees if you stimulate them too strongly. And you see that of course in certain areas of the brain that you have a not a steady response but kind of a plastic short term plasticity response and synapses. When you look at other synapses, you see different behavior. So there is versatility in the way how nerve synapses communicate the action potential train. So the way I see this is like they speak different dialects. Some synapses tend to facilitate some tend to depress during the train and this kind of code is is imprinted in the structural molecular composition of the individual synapses. And there is more and more evidence for that similar to that neuron have incredibly diverse structure and axons and dendrites fire and very, very different pieces synapses themselves are translating the action potential code into different types of dialects as reflected by this versatility of train responses. And the other thing that the synapses are facing is a signal to noise issue. So when you make a vesicle fusion competent when you make it ready for fusion, it has to be right there because you only have a fraction of a millisecond to release it. And you have to bring it into an energetic state that is kind of just waiting for this last blip of signal which is a calcium influx and that brings it into an energetic state where these vesicles can occasionally also fuse spontaneously and that would create membrane noise. Where you see an absence of stimulation you see this little blips of inward current, which are caused by this so called spent spontaneous release. And if you remove certain proteins for example that we will talk about later the snare complex that bind to the snare complex. If you remove this protein you suddenly see that the synapse faces an incredible problem of producing too much noise by sort of preventing those vesicles to not be not be released an absence of stimulation. This protein cause complex and we've been also studying a lot in the in the laboratory so so there are things to keep in mind synapse synaptic vesicles need to be kept in a certain energetic state. So that we have a maximal response when we give an action potential and minimal activity outside of this outside of this firing of presynaptic nerve terminals. And so that's this isn't kind of a waiting of functions in a nerve system you want to make the vesicle fusion efficient and their fusion properties so that they can fuse efficiently and fast, but don't make it too easy for them to fuse.  Good. So let's look at this on a on a scheme again. What does it take to make a vesicle fusion competent and ready for fusion. So we have this calcine trigger the vesicle sits right next to the membrane we call this docking. They need to be brought down to the membrane and they essentially touch the membrane. And also we have to assemble a fusion apparatus so that, for example, the forces that needs that are required to sort of break two membranes and make that two membranes into one that that these forces are available the energy for that is available. And this, this transition is can be actually triggered by the influx of calcium. Now, now the energy barrier for two membranes to fuse is extremely high. In normal cases, probably would be taking spontaneously without any of this fusion apparatus years. And that's very good because otherwise ourselves would constantly disintegrate through instability of the membrane so membranes intrinsically are very stable. And so you need a catalytic machinery that makes this transition very very easy and fast. And that's what I have been fascinating and what how can you build this machinery. So the system that we use in the lab to study is a very very reduced system. And this sort of is the stink is simplest connectivity you can think of, it's this kind of robins on cruise or situation, you have a single neuron growing on an island, and this neuron can only talk to itself, it has it forms axons it forms dendrites, you can see them here in the dendrites and blue this is so much and you see all that the axon forms, hundreds of synaptic connections onto its own dendritic tree and with this system, we can study the synaptic transmission with a single patch pipette, very easy system You know, Yuki got also very famous when she was in in Chuck Stevens laboratory she used this technology in fact to discover how synaptic attack in the council center we will meet mentioned this briefly don't is actually operating at the synapses so she showed this in knockout mice for snap the tagman using this autaptic system. And, you know, I'm also been very fond of the system because it's just very simple, very highly controllable to study the mechanisms of release. And so what we basically do is we do a simple pre short somatic depolarization of the membrane from minus 70 millivolts to zero millivolts for just two milliseconds. And then along the axon initial segment you get an unplanned action potential sort of distributing the electrical stimulus along the axon, and you're reaching all these hundreds of synapses at once, and then causing a synaptic response that you see here. The beauty of the system is the response size he see here is is quantity quantitatively dependent on factors that we can all collect in our system. We can count the number of synapses, the more synapses the larger the response, we can measure have ways of measuring the number of fusion competent vesicles that these hundreds of synapses make all together it's kind of a sum up. There's also called pool of readily vesicle releaseable vesicles, and we can just simply do that by applying hyper tonic solution to this neuron. And then we look at the transient component of the responses are all sort of these mini blip responses from individual vesicle summed up and count them essentially by looking at the charge current charge and flows and typically for these couple hundreds of synapses we have 5000 fusion competent vesicles. So let's say you have 200 versus synapses and 5000 5000 fusion competent vesicles we can make this quantification for each individual neuron. And we can put this of course in context as an optical response. And we can also look at the spontaneous release from these pool of vesicles so when we have 5000 vesicles available. And the activity of five hertz of five per second that means each fusion competent vesicles that sits here fuses spontaneously every 1000 seconds. So because rates we can directly measure the rate of release of individual vesicles both doing stimuli and spontaneous, we have means of describing the energetic state of these vesicles and quite with quite high accuracy. And that allows us to study, essentially the machinery that underlies this process in great detail and find out what are all these factors these proteins and so on are doing to make this release process possible. So, here is just to briefly mentioned how can we actually measure the efficacy of release itself from Carlson triggered responses. A classical method is to do pet pulse stimulation we give two action potentials right after each other. And then, depending on whether the second response is smaller or bigger. We know that the release probabilities high or low this is just something that has to do with supply demand calcium influx. But we can also directly measure it by quantifying the number of vesicles that are released by an action potential by the number of vesicles that are available. And we call this vesicular release probability again something nailed down to the behavior of individual vesicles. So if you have 5000 vesicles available and 500 are released here, we know that the probability of vesicle to fuse with an action potential is 10%. So it's essentially we know the property of the dice that is rolling when this action potential comes in, we say okay, an action potential makes one out of 10 vesicles to fuse. And this is very important because we can now look at different synapses and we find at the, let's say at the cortical synapse that this likelihood is only 5%, while when we look at an auditory synapses might be the probability might be 15%. And so knowing about this probability of release helps us to understand why some synapses are, you know, depressing during trains of action potentials while others are sort of showing facilitation. So the other thing that we really like to do is to manipulate the molecular content. So if you're really trying to understand how the apparatus of release itself works, we need to go after the proteins. And so for this, we've been, you know, initially, you know, as Yuki said, I started to use a lot of pharmacology to understand the synapses, develop tools to use these autapses. But then, luckily, I was also got in touch with people like Tom Sutoff on the exposure that actually made knockout mice for presynaptic proteins. And our system was perfectly designed to actually study what's wrong with my synapse if I take this protein away. And so that's kind of what we've been doing a lot over the last decades, I would almost say we use transgenic mice that lack essential components of the release machinery, and then characterize the phenotype of this loss of function. But then we can go one step further we can bring back the protein of interest using in particular antivirus we like them a lot by essentially reintroducing this protein using a virus. And then, for example, study mutations in these proteins, or the concentration of these proteins, and how it affects actually release. So the system by putting in reporters or manipulators of neuronal function here, for example, we do have a mixed culture of two different types of neurons. They have green nuclei and red synapses or red nuclear and green synapses. So you can study how neurons form connections with each other under controlled conditions. Here's an example how we use this transducer, the manipulators of electrical activity in our experiments, and brings me also to the second last technique that is important for us is electron microscopy, because synapses are functioning, but they also are structures, and the structure is obviously also involved in controlling function. So in this case, when we had a visiting scientist in this case was Eric Jorgensen came to Berlin, and they had been sort of trying to read and do the version Hoysan Rees 2.0, sort of trying to capture synaptic events on a very on a millisecond time scan using ultra electron microscopy. So the idea is to use a high pressure freezing device, a transfect neuron with a general dobson to control their firing property using optogenetics, and then essentially stimulate an action potential milliseconds. Before we freeze and arrest the structure in its native state. So then when you look at this electron microscope microgram, you see actually a fusing vesicle in the central synapse, which is something that's very, very difficult to do if you do electron microscopy in absence of syncing with the activity of the neuron. It's been extremely powerful novel technique that we've been utilizing now in the lab a lot. And the other thing that you can do is you can sort of look at these structures at the pre synaptic membrane and the position of these vesicles, and sort of ask okay this one is directly attached to the membrane because it's docked. These are the vesicles that are sort of close by. And then we give us in single action potential. And then what we see what what happens with the the number of vesicles we see. And we've what we found is that only the number of vesicles that are directly attached to the membrane is reduced in the in the profile but not the dog desert vesicle so we know for vesicle to be fusing to be able to be released it really has to attach to the membrane. And so that these are kind of tricks how you can use a system to understand it's it's it's architectural principles and and how the positioning of these structures reflect function of these vesicles in the synaptic terminal. Here is just even more advanced things that we currently trying to do is using cryo electron microscopy, which is much more native than even the high pressure reason that I showed you here we basically use no staining no no heavy metals, no fixative really membranes and trying to identify synapses why we are stimulating them electrically. And you might see this here this is a synapse and then you look at carefully to see little bumps here. And we think we don't have proof yet for this is all work in progress. We think that we can actually resolve in this technique in fusion pits and even post synaptic receptors here in quite high resolution and the long term goal of this is that we break things down beyond vesicles and membranes that we actually start seeing the proteins that underlie these activities that we've been interested in. So this brings me to that level, the protein that the core release machinery, which is going to be main focus of my data science here of my talk. These are this is so famous snare complex and the snare complex is a kind of bridging vesicle and plasma membrane.  And I just thought that this apparatus this complex is is providing the energy required for the for the energy for these men that these two membranes use which is with each other. And before fusion this so called trans snare complex because to the two different membranes are involved and after fusion it's so called system snare complex, where you fusion has been finished. And the trick here is how is the snare complex how is the snare complex operating. And how is it actually formed so these are kind of big questions in the field. It's been studied for many, many years, but we still really don't understand this in much detail. Before vesicles come up come in close contact with the membrane, the snare protein syntax and snap 25 and snap the breath and sort of float around and that separate membranes. And then you have for some syntax and being controlled is a kind of a scavenging or like a junk protein monk 18, and then somehow these membranes that inform the snare trans snare complex. Fusion complex ready for for the calcium trigger, we also have to have a calcium sensor in here. And then when calcium comes in, these apparatus allows the the few merging of these two membranes. Okay, so here's just a little bit more detail about syntax and which are mainly going to talk about today is syntax in can be in different confirm confirmations it has kind of the executive part which is a red part, and the regulatory part which is shown here in the end term is and the HPC domain, and we have here among 18 so at the start they form a complex, and then you have to come in with a vesicle snare protein them to us and after Brevin, and that sort of crawls around the snare motif. The two snare motif sort of merge here at the end terminal part. But this is only possible when these, when, when the regulatory domain of syntax and can sort of open up here. And then eventually, you have enough template that also snap 25 which is showing in green is sort of joining the the complex that the end you and ending up with this trimeric snare complex that forms over all these four bundles. And we have these at the end we end up with these complex of vesicle membrane plus membrane and snare complexes. And that's kind of the fusion ready vesicle that awaits the calcium trigger of course what is not shown here is the calcium sense itself. We don't really know where it is exactly it's also an interesting question. But one thing that we can already do in our analysis is to ask how many of those snare snare complexes do we actually need for fusion to take places a single one do we need 10. And so using our genetic tools we can address is simply by taking out to control in the concentration of one of those key proteins in this case and taxon by controlling its amount in the synapse and how do we do this. We take essentially knockouts, heterozygous mice, and then knockdown approaches so we reduce the concentration of the protein using also knockdown constructs and combine those. So essentially we do kind of a titration down of concentration of the protein in the synapse you see go gradually the signal goes down, while the synapse is maintained. And then we can look at these different concentrations how synapse is actually performed. What we find is that we look at for example the number of vesicles that are made a fusion competent is so called RRP charge drops, of course at some point to zero because we need the snares to make vesicle fusion competent. And but looking at the function and the relative concentration or relative expression, we sort of gain an idea about the, the importance of the stoichiometry of this process. And we can also do the same game by looking at vesicle release probably the likelihood of the die throwing what I told you about. And we can look at how when we constant lower the concentration of syntax and that gradually release probably drops. And from this we can be actually found that the slope of this process approximately three. And that could be indicative that essentially there is a transition between vesicles that maybe from one snare complex and multiples that cause an increase of the fusagenicity of those vesicles. And basically illustrating here that we can utilize genetic molecular manipulations to understand better the process of fusion itself. So this is just to show you the modular domains of syntax and I already told you this is a regulatory domains which are sort of opening and closing and enabling the the the business part of the snare domain to to be into interacting with the other snare proteins. And I don't want to we've been working on this quite a lot is very interesting research done here. But I want to mainly talk about to you about executing domains here the snare motif itself is the linker region between the snare motif and the transmembrane domain. This is where they form this bundle. And in between we have a region short region which is important for function and then of course the membrane anchor. So this is what I mainly want to talk to you about what we found in our molecular structural analysis of this region how that actually affects vesicles for in their fusion properties. And to show you this, we're talking about mainly this region here in the trans domain on the system system structure, this gray area here is the linker region transmembrane domain and that's a snare motif. And so the first thing that we did was to ask how important is the length of this structure how is essentially the idea is, how does the snare snare complex couple to the membranes, because something do something with they may do something with the membranes. So there's a pulling in the membranes making them sort of fuse, or making them unstable or forming kind of dimples. This is something that we don't really know yet. But we thought, let's just put a single single helical turn almost it's not even a full turn by just putting GSG into the sequence so we take the syntax and knock out mice. We made them so they are indulgence protein is gone. And then we place it by wire types and taxon or by mutant versions for example that contain additional three amino acids, either at the edge to the transfer the snare motif or to the edge to the membrane domain and then ask, what does this extension of that linker region does to release and does the position of that extension play a role in the fusion process itself. So we must say we were quite surprised to see how dramatic the effect was. In extending the lengths. I will I thought this. I expected that something will happen. But if you look at the logarithmic scale this is a release. This is the amplitude of the synaptic response, typically like five nano amp in our system and that drops by a factor of 100 to 1000. So we are inserting these three amino acids into this linker region. And so to our surprise, the biggest effect was actually putting this linker right next to the edge between the snare motif and the linker and not the linker and this And why do why was it a bit surprising because this region this linker region here has lots of basic residues, and we have the negative phospholipids in the membrane so by making this longer here by pulling this out a little bit. We moving this basic charges further away from the negatively charged phospholipids we thought this would kill the response more likely, but it was actually the other way around. What was also important. What was more important is to put extra distance between the snare motif and the man and the linker and the transmembrane domain, arguing that there must be some kind of spatial or mechanic process sort of force in place when the snare complex sort of forms zippers up up to the end and then somehow create creates force to the to to enable vesicle fusion. So we found that the fusion was essentially gone in this in this mutant. And if you put this linker in here next to the transmembrane domain, we found that the release was larger reduced and also very interesting the release was much much slower. So the release itself slowed down by a factor of about 10. This is something similar that actually Yuki found when she looked at the knockout mice of synaptic tagman so release itself became much much slower. So clearly imprinted in the structure of the snare complex is are the properties like the speed and the efficiency of the release process itself. The other thing that we found was quite surprising despite the fact that this mutant here had a way reduced evoked response and the release was slower. We found a vast increase in spontaneous release activity so the opposite of what was happening with evoked response. So we see here now that the structure of this complex can go and both can have impact in to the two main types of fusion spontaneous and triggered cuts and triggered release in opposite directions. And keep in mind we want in the synapse that evoked release is maximal and spontaneous release is minimal right so to have a big signal to noise increase signal to noise ratio. So the high spontaneous release activity is bad. And so in this tells you that intrinsically fusion is still possible, but we now actually made the release less favored, because we can't really control when fusion occurs with our calcium trigger. And so we then said okay let's look at this linker region important what what is happening in the linker region in terms of evoked and calcium triggered release and by simply making single point mutations and use basic residues to see here. So a lot of basic residues just in this very short linker. And when we made individual mutations in this residues we didn't see much of an impact in evoked release. If you make two or three or four mutations then you start seeing a bigger and bigger effect on evoked release, which is because in positive charges need to interact with negative head groups of the phospholipid membrane, but single residue changes. That's what we do here had a rather little impact on evoked response, but to our surprise, they could they depending on the position they had dramatic effect on spontaneous release. So in particular mutating the position 260 and making a reversal of the positive charge to a negative charge cost a drastic drop in spontaneous release activity, which was telling you again that this region is very very sensitive to controlling the both evoked and spontaneous release activity. So we thought to look at this a bit more careful, but one thing that Gulch who was a postdoc in my laboratory when she did these experiments made you know we typically when we use our viruses we always look for gene expression levels for the viruses, because we want to keep them all very similar. So we don't have these titration effects that I was telling you about right. If you have to low concentration then of course you get different phenotype. So what she found notice is that the band of the syntax and protein that we quantify for expression vary depending on with the mutation you saw that the vital band has the highest molecular weight. And if you have for example having mutation with a 260 we start seeing a lower band and kind of a middle band and some mutations even you see three different band sizes. And that is was a surprise nobody had noticed this before. For example if you look at Hex cells so they are non neuronal cell lines and you express syntax and they all run at a lower band. Okay, but if in neurons actually the band height is higher than in cell lines indicating that neurons make a post translation modification to the syntax and protein. Somehow altering a structure in this case we say depending on the on specific basic residues in the linker region of syntax. So that was the discovery. We thought it's really worth following up what's going on here and how is this change in band patterns related to at all to what we observe this spontaneous release activity. So, good chance sort of made up a code of of band positions going from one to six kind of arbitrary giving them a number. One is the wild type band and six hits, which has a stronger shift to the to a lower molecular weight, and she found that the band position was predominantly important for many frequency activity so a band position one gave you the lowest frequency the highest. So clearly, the post translation modification some is calling is calling very well to spontaneous release activity. So what, what's going on here. So, you know, a one obvious way of changing molecular weight is, for example, to do palmitolation of, of the protein and they are indeed in the transmembrane domain of the snare of the syntax and there are two systems, rather close to our previous lysine at position to 60 which is a very important controlling amino acid for molecular weight. And so what we did we individually mutated the systems and also and compare that the mutation of the lysine. And the result was that the white type band gradually decreased in amplitude from one mutation to double mutation and the double mutation had the similar bandwidth and then mutating this residue and doing the combination of the mutations did not further decrease the molecular weight. So this clearly indicated to us that the we have most likely modification based on the systems in the transmembrane domain that was responsible for the shift and molecular weight. We independently test this within a quite complicated biochemical experiment. Basically, you have the palmitolated residues through a steel ester, you first block all the three system groups in the protein, then you cleave the steel ester with a reducing agency. The SH group now is getting occupied by irreversible covalent bound of a biotin moiety. And then we can use this biotin moiety to be detected with strapped avidine in our western blot. And you can see here that the wild type protein has this band of palmitolated extra band that we see with strapped avidine. So this band of palmitolated extra acidity, but that's the best we could do. We're not biochemists, but we could repeat it and the signal was clear, but it's not super, super clean. And this band disappeared when we made either mutations in the lysine or when we mutated the cysteines in the transmembrane domain. So this band that we expected to see disappeared. You can also see here the shift into them in the molecular weight of syntax. So it's clear then. So from this we were convinced that palmitolation of these transmembrane domain residues are dependent on this basic residue. And so there are transfer races of palmitol groups that are sort of swimming in the membrane and they're sort of looking for motifs. One motif is of course the cysteine in the membrane, but they probably need also basic residues to actually dock on and then enable their enzymatic activity. So the question now is what does palmitolation does it directly? So we can test this by simply looking at the cysteine mutants themselves, not at this mutation and compare them to phenotypes. So again, making the cysteine mutants individually or double didn't really affect the evoked response very much. But again, we saw a drastic reduction in spontaneous release activity. So clearly palmitolation is required for maintaining high level of spontaneous release activity in these mammalian synapses. So it just brings me to the end of my first part and I will have a few minutes to finish up making this a bit more broader this finding. But essentially what we found what Gulchin found is that there are basic residues in the linker region and the linker region itself is very important for both regulating spontaneous release evoked release and release kinetics. And this residue is important for this palmitolation transferase enzyme to actually implement palmitolation fatty acid chains to the transmembrane domain of syntaxin. And having them present here affects the way how vesicles once they are primed can go infusion, they can be calcium triggered or they can be used spontaneously. And the energy barrier for going to these directions can be sort of dependent are dependent on this post translational modifications we see here. Okay, so, so that was one thing I said okay that's new model new mechanism how snare proteins are modulated by nature to have a specific function. But is this really something very specific for here or is it just a kind of a freakish thing maybe it's kind of a household function. So we noticed something looking at a different synapse system. So there are two major types of synapses in the brain. Basically releasing synapses and the chronically releasing synapse the basic release synapses the one that I told you about, they mainly respond to action potentials, the chronically releasing synapse. Essentially the release activity is dependent linearly dependent on the membrane potential of the cell. And this is something you find at the ribbon synapses in the both in the photoreceptors and the bipolar cells of the retina. So then you have essentially the member the photoreceptor measures membrane potential, and then various its release activity according to to the membrane potential so you have a light flash in the photoreceptors this causes a cell to hyper hyper polarized. And because when it's hyper polarized you have less calcium influx of calcium channel you have essentially no release. So in darkness these these ribbon synapses are highly active. Because the membrane itself is depolarized you see the membrane noise here because there's massive release activity happening chronically because the cell is chronically depolarized. So we have here two different synapses here. We have kind of a conveyor belt system where lots of vesicles kind of sort of fed onto the release sites. So these activities in this ribbon synapses, and they only stop when the cell hyper polarized is induced by light flash. And so these ribbon synapses actually use a different syntax and they use syntax and 3b, compared to the syntax and one that we find in basically releasing synapses which are sort of downstream of the photoreceptor activity. And of course, in the rest of most of the rest of the most of the brain. So, we said oh interesting so there are different syntaxes and if you look at the structure of syntax and one versus syntax and three it's isoform that is unique for synapses. At our famous site, we actually have a glutamic acid and not a lysine. And so the prediction would be that syntax and 3b is undergoing different changes in post-translational magnification and compared to syntax and one. So we thought let's look at syntax and 3b behavior in a basically releasing synapse the other way around we haven't done because we're not experts in retinal physiology. It's not trivial to measure from retina, you know light responses and so on you have to do it in the dark it's super complicated. So we stick with our simple cell culture model and studied what is syntax and 3b doing in a regular phasically releasing synapses so we made sure we can express this protein property that it enters the synapses just like the syntax and one. They go where they are supposed to go and we introduced also mutations syntax and 3b to make it like syntax and one a. Okay, so, so making the rule to make it back to lysine and so then we looked at synaptic transmission. And this is a why type evoke response for syntax and one. This is a why type response was in Texas reviews essentially gone, but if you just make the same single more point mutation at position boom, we get back synaptic transmission. And in the case of syntax and 3b, this residue is much, much more important than what we found in syntax and one is because changing this charge at least in a physics synapse is kind of a go no go in terms of release. Okay. So that was clear to us that there has been something going on in evolution, which means that syntax and one and 3b sort of diverted in their structure and then sort of mediating different types of release activity. We could also, we also took them the sister of syntax and 3b the syntax and 3a the splice variant of syntax and 3b. And in fact that splice variant is mainly in the C terminal part in this, for example, lacks the systems it's you see it here 3a has here two valence not two systems. And in fact it has at this position. It's a neutral amino acid and glutamine compared to the lysine and syntax and one. And so we gradually went back and sort of changed link region transmembrane region of syntax and 3a and thought how is it actually affecting protein structure and function. And once we put the linker region and the systems in the position, we got again a shift of the band size of syntax and 3b up indicating we reintroduce parmethylation this protein simply by putting these structures in and release was also very interesting. So syntax and three rescues, you see much smaller response and also the release is much slower. So syntax and three is not designed to be a very rapid releasing snare snare. So the fastest releasing snare is the syntax and one which you find in the basically releasing synopsis. And but we can we we make this synopsis to be reason efficiently and rapid by simply, you know, converting these region here to be syntax and one a like. Okay, so we can really play with this and understand that these difference, the syntax and three a by the way is post synaptically located. So in that post synapse of a neuron, you also have vesicle fusion with this vesicle fusion is actually important for long term potentiation. People have found this. And here we can say that whatever the syntax and three a property is, it is slightly different than what we know from syntax and one a that it is actually not very good in in phasic release. But what it does actually it shows much higher frequency of spontaneous release, compared to there's something in the syntax and three a structure that makes spontaneously is much, much higher than what we see in syntax and one a. And, and if you look at this, the the region this opens up a new air new new domain that makes us interested in spontaneous release which is this last half of the snare motif. Because if you convert syntax and three a to be like syntax and one a by making all these changes to the sea terminus, we have higher release activity compared to syntax and one a. And what is really different here is this region is is the internal part of the snare motif. So somehow in part of this snare motif, we have another region that controls spontaneous release activity. You see, you can almost endlessly study these molecules and find more and more kind of phenotypes, and that helps you ultimately understand how these snare machines actually operate, how they control spontaneous activity and and evoked release. And we even found something similar also for another isoform syntax and two, which is very, very cool to work with because it has even larger effect of spontaneous release. And again here we found that the sea terminal portion of the snare motif is critical in controlling spontaneous release because if you put simply the sea terminal portion of syntax and two into syntax and one, you get a massive increase in spontaneous release activity. And what is different between the syntax and one and syntax and two in this region are only a few amino acids. And you can just look at them they most of them conserved up but a few are different. And they look outside of the snare motif. The snare motif itself is highly charged on the surface, lots of positive and negative charges. Positive and positive negative charges means they are capable of interacting electrostatically with either membranes or proteins. And there's a famous protein that has been actually crystallized to work interact with this net complex which is again coming back to synaptotagmin that Yuki studied. And so synaptotagmin actually interacts exactly with the region that is different between syntax and one and syntax and two here. We know that synaptotagmin clamps blocks spontaneous release. So it's not only a calcium sensor for fast release, it also suppresses spontaneous release as kind of a dual function. So we now testing this hypothesis, we're just doing this now, whether this interaction that we think play a role in spontaneous release is actually mediated by interacting with synaptotagmin. And by, you know, we're in syntax and one you have these crucial to aspartic acid 231, which is mutated in syntax and two. And, you know, enabling the in syntax and one enabling the interaction syntax and one synaptotagmin, and therefore clamping spontaneous release activity. And yeah, so we are this seemed to be the case we have we make mutations in this to make it like syntax and to like and again boom you see the mini frequency goes up. And we are collaborating with people like Axel Brunge or Roseritzu which was really trying to understand this process better on kind of a molecular simulation and structural level. Okay, I'm coming to my last summary slide. Thank you for your patient I know lots of data. So, what I wanted to show you is that synaptic release has really multiple components, we see evoked release we have spontaneous release mostly the spontaneous release is undesirable because it creates noise. And but you can see that these kind of activity differs in different brain areas. Maybe in some areas, spontaneous release might be actually good thing because you don't need a physics nickel you you want to maybe have like a drippling continuous drippling of vesicle fusion happen. And so, you know, this may play a role for example and if you're looking at the eye or at the in the cortex. And these diversity and release contributes to sort of helps to shape how synapse is actually computed. Do we have a basic release activity, do we have high release probability or low release probability depression facilitation, or do we want to actually have chronically is that we need for example for ribbon synapse to convey to do its job. So, this is we know this is taking place and we really think that synapses alike in neurons, and they're firing synapse on speaking different dialects and that has some implication under the of the machinery that underlies these processes. And we know that the core proteins that are forming the release apparatus as well as accessory proteins like complexity knows not to take men are have evolved to adapt to these unique properties to make make let's say basic release very efficient or chronically release very efficient. And you can see that for example comparing different syntax and isoforms. And we think that overall understanding the mechanism, how, how this diversity of function takes place requires that we understand how these proteins also interact with other proteins. Other molecules such as phospholipids, how a fatty acid anchor that we found in syntax and one is sort of interacting with a membrane, or how calcium sensors like synaptic tag when are complex and can interact directly with this new complex. And those are the key modulators for this fusion apparatus to sort of do these jobs up here. Excuse me to the end. This is one, most of the work that I showed you was done by Gülçin Varda. She has Andrea has sort of taken over this project. And we have some older studies and syntax and John Chalra, Marie for until it did the titration experiment. Zina and Estelle are currently working on these interaction between the snack complex and is enough to tag men, and Yana has done this beautiful cry M data that I showed you. Before and we have some really key collaborators for this Erick Jorgensen introduced me to this high pressure fast, fresh and freeze technology. Jose Rizzo is a long time collaborator working particular questions of structure and function. And Tom Zutoff and these roles have been also long term collaborators working with genetic mouse models involving presynaptic proteins. And thank you for your patient. I know it was long, a lot of data, a lot of time and thank you.


Good morning. We wanted to ask you about the stabbing that occurred last night. What can you tell us? Last night at 9.55 p.m. Denver Police received a report of a large fight in parking lot F outside of Sports Authority Field at Mile High Stadium. When they arrived they found three males that were suffering from stab wounds. They had suspects that were taken into custody and they had involved vehicles which they had also cordoned off the area at that time. So a lot of people are wondering was this related to something that happened inside of the game? It doesn't appear that this was related to the outcome of the game. It doesn't appear that this was Broncos versus Chargers. It appears that this stemmed from a traffic altercation in which the suspects were backing out of a parking space and nearly struck the victim's vehicle. So how many victims did you have? And can you tell us something about their condition? We have three victims. We're happy to report this morning that the conditions have been upgraded. All three of them appear to be suffering from non-life threatening injuries. The fourth victim, there was a possibility last night there was a fourth victim suffering from a stab wound. At this time we still haven't located anybody as far as a fourth victim is concerned. So last night you reported that numerous people had been arrested for this. Can you update us on that? Three parties were taken into custody. Two of them have been released pending further investigation. One of them, Justin Manzanaris, 529 of 84, has been held on an investigation of aggravated assault for three different counts. Is it possible for us to get a mugshot of him? We're not going to be releasing the mugshot. We still have some photo lineups we're going to need to show and we need to hold that photograph until we do those. We were getting questions last night that this might be gang related. Can you answer anything about that? Yeah, we don't have any indication that this was either gang related or gang motivated. Sergeant, if we wanted to get more information or if you have more information, how would we get that? As always, you can follow us at Twitter or Denver Police at Denver Police. And thanks for watching Denver Police News.


Welcome back. We have our guest standing by for a first major conversation right here on the breakfast on Plus TV Africa. I hear as federal government through the National University's commission yesterday afternoon we drew it secular which ordered device chancellors, pro chancellors and governing councils to reopening or reopen rather the federal universities across the country. Earlier secular was addressed to all the VCs, the pro chancellors and chairman of governing councils of federal universities ordering them to reopen the university. So the second secular which was also signed like the first one by the director finance and account of the National University's commission, Samo Nasi, indicated the commission was withdrawing the first order. Don't did not explain why. The letter tagged withdrawal of secular, it's cited in the number of that secular which is quite long I won't go into it. Dated September 23, 2022 partly read, I have been directed to withdraw the NUC secular reference number indicated. Dated September 23, 2022 on the above subject. Consequently the said secular stands withdrawn or pro chancellors and chairman of governing councils as well as vice chancellors of federal universities are to please note further development and information would be communicated to all relevant stakeholders. Please accept the assurances of the executive secretary's warmest regards. Joining us to discuss what can be termed a day of a ding dong affair. If you want to put it that way is a lecturer, senior lecturer and political scientist from the Bayero University counter professor. Sanif Aga is great to have you back on the breakfast. Thank you. You've been in the education sector in Nigeria for a while now, hence you're with us in a successful career as a professor.  Yeah, professor, can you hear me please? Yes, yes, I mean you've been around for quite some time in the academia. Have you seen anything like this before? Yeah, you see, yeah, I've been in the profession for over 40 years. And throughout my stay, I think I have seen something similar to this, like the time of Brengida when Asu was banned and we were ejected out of the university campus. And so in terms of draconian measures, I can say, yes, we have seen similar things over time. But in terms of confusion, you know, given Oda withdrawing it, I think this is the past time we are experiencing it. What would you say, you know, the problem is at this point? Is it that we don't understand the dynamics of education? What exactly could he be? Is it really true that, you know, we don't, we don't really have the resources, because that's what the government is saying. And I mean, the questions are almost endless, but I want you to unsign this slide. Are the concerns of Asu not valid? I mean, the issues that are asking for, is it rational? What exactly is going on?  Yeah, you see, I believe it is rational because over the time we have seen how the system deteriorated to such a level that we sometimes conduct lectures even in open spaces. And I think this is not good for the country, because the country cannot develop above and it's on educational level. So I think what Asu is asking is not to go into the hands of the electorate, but to, you know, inject it into the system. And the government, which is saying that it doesn't have resources. But at the same time, you know, it budgeted 6 trillion Naira for oil subsidy, which is, you know, a bog of things. And the other thing, if you look at it over the years and what the government is trying to do now, it has spent over 16 trillion supporting business people. So if you can just inject less than 10% of what you are injecting to private sector, I think you can go over this problem. But even the government, you know, I say there is no money and it is a lot of money in other sectors. I don't see Asu will even raise an eyebrow because they will believe that it is a genocide. But here you are, the government is pumping lots of money more than 10 times what Asu is asking into the system. And you are still telling the people that you don't have money to inject into the educational sector. And you know, educational sector, what Asu is talking about is about the common mind making it affordable for the poor people. All right. What do you think is responsible for the U-turn? I think there are two issues to me. One, I think is the legal dimension. You know, the government erroneously or rather wrongly took the case to courts, industrial courts, and now Asu has appealed. And the government sees that if it goes ahead with the whole issues, it may be it will be seen as an attempt to do things out of the law. That is one. Secondly, I think the government has started, you know, listening to opinions that it will be too dangerous for the government to open the universities and to put the students together in place. So, you know, you are not igniting a time bomb, putting the students in one place, and you are starting containing tomorrow and then doing nothing, I think that will be too dangerous. So to my own thinking, I think these are the major reasons why the government decided to make the U-turn. But do you also think that this is connected to the fact that the government, I mean, if you look at it, campaigns will be starting tomorrow. And this might also be connected to having some sort of political gains. Do you think that there's any connection with the order that's been put out, reversing that order of the U-turn, with the fact that campaigns are just starting? Okay. No, I think that the idea to reopen was to scorch a political point, that they are doing something about it after starving with their masks. But now, I think they are seeing the possible consequences of getting the students into the universities and nothing is taking place. So, you know, the anger will mount higher, and you don't know what is going to happen. So I believe perhaps the government might have listened to, like what the committees of the Vice Chancellor said, and perhaps I don't know whether they might have gotten stupidly reported that it will be too dangerous to allow the students in one place sitting idle. All right. In your opinion, the legal aspect of this, you know, with a lecturer saying that they will not go back to the classroom, when such an order is issued by, is given to the NUC, is NUC an independent organization which is able to say we on our own are issuing the orders? Do they take directives from the Minister of Education? Because I know that, for instance, for any university to be approved, for courses in universities to be approved, the NUC embarks on inspection visits, you know, to determine what should be done, you know, they would say, okay, we need more doctors or people with a PhD in the lecturing department of this, this is the faculty, for instance. Does the NUC take directives on what to do and how to carry out its activities from the Minister of Education? Oh, yes, it's true. You know, NUC is the Ministry of Education and, you know, the leadership now of NUC is working hand in glove with the Minister of Education. So even if it is on a personal basis, they will do that one. And constitutionally and legally, they are under the education of directives from the Ministry. I think that by the NUC, you know, took what it did on the issue of the two circulars, because even if they didn't come out to say that they were directed, but I think it's a fact that NUC cannot issue directives without the directing of the Minister. And look at it, they just put a director and say, okay, he's the one who's reopening, I direct him, the university will open, and he's the one who's saying that the university should not be reopened. So I think from that one, you see clearly even what he said that he was directed. I don't think it was those, the NUC secretary that directed him. It was from above that he was directed. Well, what do you make of the elections? I mean, the elections are almost here, we're talking about 2023. And the schools are still shot down. I mean, students have not returned to the classroom. So how many more months again? It might just look like, this might continue for a long time. Do you think that this would have an implication on the elections, especially the choice of who the people would actually cast their vote for? Yeah, I think it will have. But I don't believe that it is going to last long because for political reasons, you see that the politicians will come out to do something so that they can score chief political points. Already, I think a middle ground has reached, reached, but everybody is, or each side is trying to change, you know, its models. It is all models so that they cannot be seen as succumbing to the other. But once the elections are all started, I mean, the campaign started, so I believe some politicians will take the initiative so that they can score chief political points because they know it will be dangerous and disastrous for them to keep the students for long to the election period at home because they don't know what will be the consequence of that. Okay, Prof. So we hear that, of course, now let's imagine for a second, I assume that the second secular was not issued. When the pro-chancellors, vice-chancellors, chairman of governing councils meet and then open up the universities, how are they able, would they be able to compel the lecturers to go back to the classroom? Since most of them have been saying that they will not teach, they will continue to do their research from home and like one lecturer from the University of Abaddon said, conduct their community service from home. Is there a way for the university establishment administrators to compel the lecturers if that first circular was what was held on to? Is there a way they would have been able to compel the lecturers to return to the classroom? I don't think they have the support. They can try to take their draconian measures and say, okay, we are not going to pay you, you must come, then you must sign register and so on. But I think doing that will be counterproductive because what Asu is saying is that if the salaries that we sell are paid, they will not work hard to compensate they missed. Now, if you post them to go, you are now breaking, hello, you cannot tell somebody to go and work, you didn't pay him. So at the end of it, the student will be the victims. Even if the lecturers decided to go and work, they will not continue with what they missed, which means people are going to automatically repeat and those who are admitted newly cannot be admitted. I mean, so much seems that is there. So I think the government is aware of all these things, that is why they are trying to get, you know, landing ground for the system to go. That is number one. Number two, if you look at it, technically, the universities are not close. It's only that the lecturers are not going to take them place, but, you know, administration continues. So when you say, go and open, they haven't closed, so are they going to open? So I think that some of the issues that maybe the policymakers look at and try to see a safer way to get around the issue. Professor Sane, do you think that that's a solution to ending the strike action when it comes to us? I mean, having lecturers every other time lecturing the students, is there a possible solution to ending the strike actions by lecturers? Yeah, I think there is. Already, you see, since they reached the agreement of 2009, and, you know, the government has memorandum of understanding, memorandum of action, and so on. Had it been the government is sincere in implementing these things, I think, we'll be able to head of strikes like that. Because what they are asking is, something such amount of money is a long-term plan. You are not going to put it this year. You can start to implement it gradually, like what a former president would like to eat by putting about 200 billion, I think. So over the years, they could have done and reached that place. But when the government decided to say they are not going to do it, I think that is well by problem. And the other thing is, if you look at it virtually, everything that Assad tried to do is to try to conform within the laws, okay? Even on strike, they didn't go off outwardly like that. They started by giving notice and doing this and so on, so that at least they study the laws and take cover in that one. If not for that one, I think they could have been crushed easily on legal ground. So I think if the government could come down, you know, Assad will shift a little, they will reach in the middle, and the issue of strike will be as soon as possible. Some people suspect that the second circular was issued as a result of probably some realization that there is an application by the union, ASU, represented by Fermi Falano Essein, who is a legal counsel to ASU in this matter. There's an appeal before I think both the National Industrial Court, because that's the Court of Origination, original trial, and then the Court of Appeal in Nambuja over this interlocutory injunction order made by the National Industrial Court. So some suspect that the second circular could be because of that realization that there's an appeal which was accompanied by a request for a state of execution, pending the determination of that appeal on notice. If that state of execution request is denied ASU by the Court of Appeal, whilst the appeal is going on, if that state of execution request is denied ASU, meaning that the order from the National Industrial Court still holds, would you expect the lecturers to return to work in the Middle East? I told you that what ASU tries to do is to work within the laws. So if the laws say that they have to go, I mean the appeal concluded that they have to go, I think they will go. But what is likely going to happen is that the issue of no work, no pay, will still stand, because legally you cannot expect somebody to work while you don't pay him. So at the end of the year, we are going to have another crisis. They will open and they will go, but they will not make up what they missed, and they will say let's start from a clean slate. And that will be too dangerous for the system. I have to go now. Thank you so much, Professor Sani Fagi. We appreciate your time. Thank you very much. Professor Sani Fagi is a lecturer and political scientist at Bayero University in Kano. We appreciate you always and as we approach the political season, we hope that we have your time to share your thoughts. We'll take a breakdown. When we return, it will be time for us to look at the second conversation with the issue of the national group collapse. Please stay with us.


Quick quickly about me. You just heard it, but I built these things and recently also initiated the ESE 20 token standard with italic and That became a lot bigger than expected Yeah, so when we think about identity and this talk is about identity and it's also an ESE What is identity, right? So if you think about identity today My clicker doesn't work Like it could be for example your physical Attributions right like it could be how you look or your biometric data and some kind of like physical properties attached to your physical person and Your body for example, right? But an identity could also be the group you belong to like the country you belong to or even the history over time Like the reputation over time or the things you experience or your ancestors experience and so on and this is all what we kind of aggregate to our identity Today obviously identity are mostly like some kind of like forms and certifications We get from our government or we get from like all kind of other places For example, we have social security numbers. We have plastic cards We have passports and so on This kind of like is what we think our identity is today and this is what we use in the world to represent ourselves So when we think about a passport for example, what is it really right? What is a passport? It's basically the fact that there was a government agency Bureau grad sitting somewhere and I came along with a birth certificate and a photo and Maybe they measured my biometric data and they figured out. Yeah makes sense It's all matching. I give you a passport So basically it's a collection of Things and what it is is actually collection of claims Because what I'm having here is I have a claim The birth certificate which somebody gave me when I was born, right? It's already a claim, but that person who was there maybe was the doctor or maybe was the hospital That I was born there Then I take a physical claim which is basically a photocopy or maybe some biometric data scanned and Then it creates a new claim now done by this guy here There's no laser pointer there and This creates now this new claim a passport So basically almost everything It's just a lot of like attestation we get from each other and we give to each other right for example We could look at here this use case. I have passports or I get driver's licenses Here or for some life a university degree and this is in the first place issued by a government to a person Who can issue that degrees to in universities itself and they then issue a degree and this is really hard here To do another person, right? So it's a lot of claims on the end. It's all claims It's a humongous network of claims and we came up with a lot of different type of claims degree certificates and whatnot and we collect them over time and it's kind of like how we represent ourselves in the world Maybe not necessarily with our friends with our friends. We might use social networks, which is also kind of like a collection of claims But if we go do any kind of like Company cooperation, we make an interview or whatever we just take all of these claims You know we show them here look at all of these titles and I did this and this and that and This basically tells the other person a yeah, I trust this university. It's a big one looks like a real thing here what he has Here I might just hire him for example, right? The problem with the current systems is that we have this huge institutions which issue claims and I'm the tiny little guy and I will receive these claims and I Kind of like the power sits in these institutions and it sits in the humongous data walls, right? so identity claims today is basically Not necessarily with me. I mean it's with the papers I carry around but it's kind of like a more like a monopolized ecosystem What if we could decentralize all the things we could just take that and Distribute that power and that ability to claim things about each other and Where do I have to point I have to put this way So for example when we think about Uh Standards right so what are standards good for and I initiated together with Vitalik? Basically I proposed that standard and it was a huge debate and on the end everybody started like using that standard It really helped foster a few things for example if you look at the token example, right the token standard and how it kind of Came to be is we had already things like Bitcoin and then people came with this idea. Hey made let's make colored coins and Then there was this project master coin out of it later Splitted off counterparty and what's the idea is that we can basically create coins on top of Bitcoin We have the same security like the like the Bitcoin network But we can create new coins and then companies suddenly can issue their own coins And that's what's all really nice and neat and it was kind of used but not really big then a theorem came along and Also people started issuing tokens Inventing their own ideas of how a token can look like and we had things like digix the gold token Which was kind of like the first at least I know of rather bigger custom token But what really actually made this very interesting and very powerful is When you come up with a standard like if you say hey a simple set of rules you basically say hey Let's make these three four or five functions Let's all use these function as the Operator the interface towards the token and suddenly we can have wallets exchanges other contracts interacting with those tokens and Suddenly you have a complete mess Yeah Yeah, I mean this is actually what spurred all this this craziness let's say Because suddenly we have a rather simple standard which doesn't really even determine much But it allows so much flexibility still inside that all of this experimentation and all of this crazy new ways of how using it became possible and that's why it took off so big and it's still like Obviously in the rocket ship flying wherever So It's actually all about standards if we have standards it allows us to basically interoperate and be compatible and especially in a platform like Ethereum where you have an endless amount of possibilities of what you can do and how you can interact and what you can create Standards are really the key to make a viable ecosystem flourish So I thought about like a few years about Two years about like kind of like how I then you could look like was working back then With a client and thinking about this idea and I kind of like had this idea Okay, it should be a smart contract and that should represent you and la la la la and I thought about it and really until now I didn't came up with any way different solution than the one I had back then and Actually always expecting that somebody else would came up with a standard and Megan ERC which didn't happen. So I did it So I created this standard or initiated it And on the end it will only become a standard that everybody Participates in everybody agrees, but that's the whole point of a standard and it now has the number seven two five And I created this this standard which basically defines a smart contract which represents you I don't represent not even a person. It could also represent a machine or even could represent a an item an object or a company So this identity standard basically Consists out of three components. So it itself is a smart contract Which is representing you and it has these three components. It has keys, which are your operators It can execute so you can actually act on the blockchain as you and it has claims So I will now explain these three pieces here and then show some examples of how this can work The idea is basically that you have these keys and this is how you act in the world, right? So for example, you can make transactions obviously on the blockchain You can send money around and then you can Prove with that that if this is one of your keys in your identity that it obviously must be done by you I Can also sign any kind of documents or I can send sign any kind of proofs or messages in the real world Proving that it was me doing that as long as I attach my identity to it The person seeing the signature can verify the signature and can assume a very high likelihood that it was me So actually the list is like unlimited because what could do with these kind of things It's like not very limited in this sense. The idea is and this is the beauty of Public-private key topography is that we can have these kind of control over we can basically like validate ourselves In the real world and others can provably Validate this this as well. They can check that this is correct without ever I ever have to reveal my password or my secrets or whatever and I think we are just getting started into the age of Key signing right so will we do this and all the time for basically everything and a really simple use case for this for example would be logins, right? I can simply Sign the message I want to go into your website and they can prove that maybe even check my identity and Do some kind of verification process and they can let me in and I never send my password over the wire Or anything and I could even use a third-party to factor the cation device the other part is and Because it's a smart contract. I can execute so this standard has a execute function Which allows you to act as yourself on the blockchain so you could either use the keys to do actions on the blockchain and then somehow like Point the people you interact with to your identity so they can check things about you or you can actually act directly through your contract and It's really important thing is that you can add other claims to other Identities or to other people objects or even other smart contracts, which might just be code and You can claim that for example You verified them or maybe you're an issuer about claims Maybe your government and you can attach claims and it's very approbably coming from that identity and The third part and that's actually the important part We're just its own ERC After some discussion we decided moving it out in ERC 735 and Basically making it a substandard of the 725 and the idea is really to have a way to attach claims to a smart contract and now we have all to debate exactly how this can look like and I have an initial draft up there of how this can look like and how it can work the benefit of claims in Your own identity it can be added by anyone, but I always have to approve it right if it's my identity I want to need to I want to be able to approve it because I don't want that things are attached to my identity Which I don't really agree to this is especially important if I'm a person if I'm a smart contract Maybe an approval is not even needed.  So basically The person who issued to claim about me on my identity once I approved it They are technically in control because they issued it, right? But if I don't for example, if they change something about the claim and I'm not agreeing I could always remove it Because it's on my identity so I can allow it to add but I can also take it off anytime if I disagree with a change Or for example if the change to data hash and I never really was in their office again or whatever and the important thing about the claim structure in this standard is that the Claim itself contains in actual signature and The beauty about having it index mark on it and having a signature that this could be any kind of signature The difference here for example, if you would make a registry of claims, then it's always the always have to be an ethereum Signature, it's an ethereum transaction and the smart contract registry will determine how these claims are added But this is a really open container because it can be all kind of different type of signatures in the future doesn't need to be an ethereum one. It could be anything and It obviously contains the actual claim reference or claim data It really depends on what kind of claim it is Most of the cases you don't want to store any kind of actual data on the blockchain You don't want to store an actual claim like it. Maybe here your The bytes of your photo or maybe your actual address you only want to claim at the claim with the reference To the actual data. So the actual data will still sit somewhere else and this is to be debated This could be sitting in swarm IPFS or this could be sitting with the company directly still But in some cases even it might be you want to have a bit mask or something which actually says something about you Right in the bytes of the claim where there's more detailed Claim detailed so to say in it So basically how this could look like in detail how it's currently in the in the standard and because it's a smart contract standard It's obviously code We have a claim type and the claim type for example says Maybe it's a claim type zero one two three four five Maybe we say zero is the claim for a biometric data and one could be the claim for an address So if I see you on your smart contract, you have a claim type zero and a claim type one and They are both issued by an issue. I trust Maybe it's the German government and maybe it's the Deutsche Post which knows your address and I trust them Then the only thing I have to check is does the signature actually contains Contains actually the holder address which is you at the address of the of me in this case It contains this number here the claim type and it contains this little data And it comes out to an address which is still owned by the Deutsche Post or the German state Then I know that claim was actually issued by the Deutsche Post or the German state and as long as any external party trust that issue on He can be completely confident that you have an address and that you are human or Maybe some kind of flesh thing. I if you have biometric data and That might be fully enough to allow you access to something to issue credit or to issue an account or whatever you want to do in an automated process so it's all about Basically trusting the claim issuers and it's obviously that depends on what kind of like issues This could be and in the beginning this probably will be big institutions But in the future this could also be peer-to-peer networks or other kind of reputation systems or whatever the claim Can be it's it's completely open, right? So how would the process look like and I have three minutes I have to go faster for example like a off-chain example could be here I'm walking around with my Smartphone simple use case I want to get into a disco and The guy has to prove that I'm over 18 or whatever So he would basically send me from his phone a random string and this random string is something I can sign and I will sign this random string plus my identity address and send it back to him He will be able to verify Okay, this was signed by this key and he signed my random string So obviously he could he had the private key he could sign it properly and yes, there is his address his identity So he goes check on the blockchain gets the claim he needs or whatever claim that is right Maybe it's the age whatever or an age certificate. That's over 18 says that says over 18 or so He basically checks the claim and this means he checks the signature and And then goes and checks on the smart connect of the issue that actually the key he's designed with is still there So this would be the power for the issue to remove a claim even though that I might not agree to so as long as the signature is correct and Returns a address still owned by the issuer this person here The bouncer can be confident that this person is whatever the claim says it does And it might be a very abstract information. Maybe just over 18. It doesn't even mean he's 20 30 50 or 110 it just says it's over 18 or He's allowed to just Go in for whatever reason, right? So you you could do this in the off-chain world basically by going around and signing messages all the time And then people can always go to your blockchain identity and check Hey, who is that person does he have the right requirements and you can verify these kind of requirements? If I do this on the blockchain, I Basically can act through my identity. I can call a function on another smart contract and this could be the super important Cool file smart contract and I really want to use that function But this function requires to check some identity stuff. Otherwise it can't be used So the smart corner will automatically realize, okay an identity is calling me So I get the claim from this identity smart contract back. I will verify the signature right in the smart contract Go to the issuers smart corner and check that key is still there. I know the key is still there So it's a valid claim still attached here. I trust the issuer I don't even need to know the actual data most of the time. We don't care about the actual data I don't know care how what your name is or how you look like until As long as I know that you are a person and that you have maybe a residence somewhere where it's Where maybe it's the country or whatever or maybe it's just enough to know that you haven't residence at all And that's it. I can continue with the execution of the super cool function in this case so you couldn't do this you can use this on chain and you can use it off chain and my time is off and You can basically do these really simple steps of Actually two or three steps to completely verify the claim and it's attached to my identity. So I'm full in control and This allows basically all of these interesting things and it combines these these two little rather simple standards And it gives enough flexibility that all kind of experimentation can happen So we can figure out what kind of claims we trust how these claims can look like in detail Where they are stored? How could you even verify the claims actually content may be in a CK snark proof outside of the blockchain? People can come up with all kind of systems. They want to Claim about each other and as long as we follow the simple standard we have that interoperability Which can create all of this network effect we see with ESC 20 for example so it's all about discussing and Coming to an agreement what makes sense start playing around and making a huge experimentation around it you


In the 1940s and 50s, the American Medical Association was not only saying that smoking in moderation was not a problem. On balance, smoking may even be beneficial. After all, most physicians themselves smoked, though, so how bad could it be? When the American Medical Association is saying that, where could you turn back then if you just wanted the facts? What's the new data advanced by science? Well, she was too tired for fun, and then she smoked a camel. Babe Ruth spoke of proof-positive medical science, that is when he still could speak before he died of throat cancer. Now, some of the science did leak out, causing a dip from about 11 cigarettes a day per person, on average, down to 10, but those that got scared could always choose the cigarette that takes the fear out of smoking, or even better, choose the cigarette that gives you the greatest health protection. Now, if by some miracle there was a smokingfacts.org website back then that could deliver the science directly to the people, bypassing commercially-corruptible institutional filters, you would have become aware of studies like this, an Adventist study out of California in 1958, that showed that non-smokers have at least 90% less lung cancer than smokers. Now, with so much money and personal habit at stake, there will always be dissenters, but look, given the seriousness of these diseases and some total of evidence, we shouldn't wait to put preventive measures in place. So, if you were a smoker back in the 50s in the know, what do you do? With access to the science, you realize the best available balance of evidence suggests your smoking habit, probably not good for you.  If you wait until your physician tells you between puffs to quit, you could have cancer by then. If you wait until the powers that be officially recognized, like the Surgeon General did in the subsequent decade, you could be dead by then. It took more than 7,000 studies in the deaths of countless smokers before the first Surgeon General's report against smoking was released in the 60s. You think maybe after the first 6,000 studies, maybe they could give people a little heads up or something? One win is how many people are currently suffering needlessly from dietary diseases. Maybe we should have stopped smoking after the 700th study like this. Now, let's fast forward 55 years and know there's a new Adventist study out of California warning Americans about something else. They may be putting in their mouth, and it's not just one study. According to a recent review, some total of evidence suggests mortality from all causes put together. Many of our dreaded diseases, ischemic heart disease, circulatory cerebrovascular diseases like stroke significantly lower among those eating more plant-based diets, in addition to less cancer, diabetes. So instead of someone going along with America's smoking habits in the 50s, imagine you or someone you know going along with America's eating habits in the present day. What do you do? With access to the science, you realize the best available balance of evidence suggests eating habits are probably not so good for you. So do you change or do you wait? If you wait until your doctor tells you between bites to change, it could be too late. Just like most doctors smoke back then, despite the overwhelming evidence published for decades, most doctors today continue to eat foods that are contributing to our epidemics of dietary disease.