Welcome to MOOC course on Introduction to Proteogenomics. Today we have scientist Dr. Suman Thakur from Center for Cellular and Molecular Biology, CCMB Hyderabad. Dr. Thakur will talk to you about mass spectrometry based quantitative proteomics and how it can help the cancer research. We will also talk about why quantitative proteomics is very important, in which way various tools available for doing quantitative proteomics have helped to understand different diseases. Dr. Thakur will focus on why we are not getting clinically relevant biomarkers for all the cancers and why one drug cannot cure all the cancers. He will talk about different specifications for mass spectrometry based columns, for example the gradients, the column length, another type of mass spectrometry parameters, how it could be optimized to obtain good results. He will also talk about why the number of proteins decreases in labelled analysis as compared to the labelled free quantitative analysis. He will then talk about anti-cancer compound screening and how the proteomics cell biology and animal studies together could only help to find out the clinically relevant information. So, let us welcome Dr. Suman Thakur for today's lecture. Mass spectrometry based quantitative proteomics in cancer. So, anyhow you are well trained and thanks to all the previous speaker who has set the stage and I should just move it. So, all you know proteogenomics, so genomics is here, proteomics is here, middle transcriptomics, metabolomics for us these all are different subject, but for body all are happening, billion year trillion year of evolution has made this. So, this is not a one day anything, maybe this is new subject for us, but what is here? So, biology change, thousand years before also question what is life, today is also question what is life, most probably thousand years later will be also question what is life, what is evolution, but what change that you have to understand, technology. So, technology drives biology, you get new technology in your hand and you try to answer different question, if you want to do something in life you have to ask question. What is the question? Find a specific anti-cancer compound for particular cancer without side effects, is it possible to find a drug without side effects that is the reason you ask question what is not easily possible. Second, find earliest stage and cancer biomarker, cancer is such a word till you discuss in symposium and lecture hall it is fine, if you see any patient your near and dear you will be second very badly. So, that is a bad word is a cancer basically is a disease, still after so many years hundred years of research we are not able to cure it, we are still in the middle, so what you need earliest stage, every doctor ask earliest stage, but what is the question common cause there are different organs in the body different types of cancer are there, if all are there then cancer should be common, this question you can ask and we try to ask this question and I thought take select eleven or ten cancer cell line, it is very nice to we have to get clear this is the cell line, same thing we are trying to repeat on the tissue also.  So, different type of cancer are there, why not one drug is there for all the cancer, impossible, origin is different, metastatic is different, so that means it is very complex, but then you have to take if you want to do anything in the proteomics, you have to select one control and this is non-cancerous cell line. So, we thought we will take this ten cancerous cell line and very childish way you think you get 5000 protein, so then in L10 or L11 you will have almost whole human proteome practically it is not possible or you do different-different fractionation and you come to 10000 protein in each cell line, still you will not able to cross 11000 maximum 13000, so what is the matter in that means all proteins in all the cancer are same only quantitation is getting changed, some place someone is expressed, some place someone is expressed. So, that means what expression level, quantitation, so finally what came, quantitation is important in the experimental and it is happening inside the body, but how you will do this, this is very simple, we want cancer full proteome that is one or few proteins protein chemistry, you will see some lab is whole 20 years has been focus on one protein, but some lab will not like one protein, why one protein we should have 1000 protein, but what you will do is 1000 proteins, again what is miss whatever is your interest that you have miss, so finally if you want to make a drug or target you will target on one protein, so what protein mix is giving you, completely back cycle to find the target and come back on the one protein and make the drug, same things we are trying also to do, but before that I will go with little bit problem is there, there is nothing everything is good good good in life, but mostly we talk good good good thing, if you go to say the biomarker things, different disease same marker, same disease different marker, all you will get confused and you will end up with the nothing, so bottom line what we need, need unique biomarker, that is the whole thing and field should grow it or maybe you need set of biomarker, just like if someone is telling no no this is this disease, then again you go to cross check, you go to verification that is thing has to be done, why all these things is happening, answer is here billion year of evolution you cannot easily search find in one day directly sought it, it has to be systematic study or if you are by luck you are hitting in the dark it got hit, both are happening together and both example I will show you how we fail, how we pass, we thought instead of 15 centimeter make half a meter column, think little bit weird everyone will tell you crazy it works you are best, you did not works you still carry with the tag crazy, so now 15 centimeter, 50 centimeter column it is long column make beat side everyone using 5 micrometer, 3 micrometer, half what will happen HPLC pressure will be very increase, to increase the pressure simple use school knowledge Charles law, Boyle's law, heat it down the temperature down that increase the temperature down the pressure and somehow you will manage with old HPLC this 50 centimeter column long run and 5000 protein came that time in one single shot this was just example same technique we have developed now recently we have published how long gradient we can use and after 12 hour if you are using the gradient recently published in 2018 there is no use, so it is saturated, so need of fixation how long gradient, how long this and that. Now this technique you should use in silak that I am going to tell about the cancerous cell, so this is just introduction now I show you label free quantitation 5000 protein comes in one run, but when you do silak you get less protein why because now 2 peak is needed to identify and quantify a protein, so you increase the complexity if you increase the complexity sensitivity has to be compromised or your protein number has to be compromised but we know all the way how to increase the number also what if in one hour you got 3700 make triplicate now and then quantify here bioinformatics you like or dislike bioinformatics you have to use, so now this is the place 1 into 8 hour you got 3000 now run triplicate you get 4222 because which one peak has come in this run another peak came in this run by bioinformatics you merge and you can increase that, so this way you are showing, but there is the way triplicate 4 5 times 6 times how many time this is showing almost in triplicate on 4 almost you are getting close, so there is no need of going to unnecessary too much you have to keep your temptation and limitation that is also very important otherwise you are doing only one thing, now same thing we did with all this 11 cell line and then we try to find is there any common thing is there or we have to stop on 11 first we did 1 then gone 5 then 11 now we are planning to have 25 or 50 cell line to get complete idea, but we cannot decide in one it is very costly affair and we have selected in the broad range, now by this if you do deep proteome you will get 10000 protein in each cell line by fractionation, but we thought we should go single thought also and we should see in one run there is no comparison between any fractionation 4 fractions and unite no bioinformatics one run direct result and see that both approach we have used now you see what happens here it is single thought one thought whatever came this is CID silent you know CID, HCD you would have heard high energy today is the era of HCD. Now you take hack 293 light plus heavy so hack is there light and heavy now you are comparing that there should be theoretically any difference in quantitation up regulation down regulation it should not that is the reason when you quantify identify 4200 quantified 91 percent in 4, but when you did in 1 I told you already concept here you quantified only 78 percent here 89 percent here 91 percent. So what happens when you do multiple run you are going to quantify more, but now I am increasing 5 cell line I am mixing 5 cell line together and then I am trying to see what is happening one thing you see if one cell line give me 4,299 protein mind tells 5 cell line will give more correct or not simple thinking do not think too much answer is no then we thought 11 cell line will give more answer is again no see this pattern is containing single run is giving less triplicate is giving more and here you are getting more, but when you are increasing the cell line nothing is changing that shows your technology has limitation of course 5 cell line when you put you have more protein 11 cell line when you put you have more protein, but technology has limitation that means still our so called well developed well costly mass spec need to develop less or more very more so still price will increase a development will continue. Now see now this is same CID here compare hack 293 versus hack 293 so there is theoretically there is no difference when no difference so what happen anyhow in quantitation we do not take less than 2 fold ok see majority of things is falling in this less than 2 fold and whatever it is showing 1 percent that is almost a error ok. Now I have taken 5 cell line in 5 cell line I am taking HeLa versus Jurgat HeLa HEPG to GamG FCF7 so this HeLa is inside that 5 cell line, but I am able to quantify and see 2 fold that is falling only 84 percent that means others are changing ok here is the very less change here you can see more change that means different cell line has different things correct. Now here I took 11 cell line it is coming almost same that means because my technology is also coming close and result is also coming close so I cannot comment on that clear. But that shows me 5 cell line has more protein or different protein compared to that and if theoretically same 2 cell line if you compare where is the up regulation down regulation if here there you get up regulation down regulation that means better to stop the experiment ok. So that shows your control things are going in right direction, but this HeLa is inside the 5 cell line and 11 cell line with this you plot you have to learn all the R and all these things today by plot you can see the 5 cell line is not that much sharp 11 cell line is more that much sharp that means quantitation is better when you have multiple cell line a chances to find that protein is very high same things you do with HCD 1, 5, 11 cell line and you will get different type of things and we are fine here, but this HeLa is again inside that. Now HeLa compare with Jurgat you will get only 70% no changes almost, but HeLa with 5 cell line HeLa with 11 cell line you are getting more to cover it. Now it is different you are comparing again you see that HeLa with Jurgat your line your graph is not density plot is not so sharp here it is better and 11 cell line better. So this is giving me indication that my 11 cell line are higher stock I will able to cover more protein of that if that mix is ready that proper things is ready that means I will have more chance to quantify because in human I cannot label HeLa ok it is only possible in cell culture ok. Now you see here here we have taken one cell line HCC 1, 5, 9, 9 breast cancer this is not mix present in the mix of 5 and 11 now you see here chances of getting quantification is more or less because it is not inside you will get different things. So this concept is coming clearly how you make the mix we do we need a master mix do you need a this and whole analysis idea is still we are doing that what common protein is there in all the type of cancer where it is getting click or where it is getting a start that is still under the way most probably next time I will show you more and better same things I am trying also to go with the tissue now when 11 cell line we can take now we know what is going to happen now 11 tissue or 20 tissue we are trying to do all cancerous tissue and we are looking what is the common and can we find one common place where all this get trigger that will help will we do not know what will happen with this you again see this density plot always 11 cell line is coming better ok you all have learned R and with this I will move towards the little bit cancer drug discovery that is the my favorite hard work of VHD student and it is somehow giving good result. So what is there development of anti-cancer compound use cell biology mouse model and human model you have to go finally clinical tried so that is the what simple cancer means tumor break the cluster induce cell death apoptosis and reduce cancer cell proliferation three things as a target different compound we have a screen there you would have heard a company a screen 10,000, 15,000 compound library and then come to one target we thought how can we do a smartly here this is the end childhood cancer mostly happen in childhood time before 5 year why it happens it is very tough to tell no one knows it can happen in one eye it can happen in 2 eye and both the eye can be affected there is the drug with very high side effect carboplatin, etopocyte and vincristin mostly natural towards ok so we thought nature may be have something with this now how to think about the cancer how much unfortunately this retinoblastoma is very high in India or developing country India, Africa and all this why there is no answer of that but if you look all these things you will see 10% of pediatric patients have retinoblastoma ok in that also highest incidence of retinoblastoma is in Africa and India why no one knows this is the cancer related with the gene because you are in proteogenomics and this cancer is related with the gene retinoblastoma RB you would have heard cancer suppressor gene this is directly linked with that ok so this is sometime hereditary sometime environmental but there sometime no one understand why it happen ok now people are still thinking which place it is just getting a start and recently one in 2014 paper came that they tell it is mostly a starting with the cone precursor cell ok so this is the little bit evidence has come so this is below 14 years is non hereditary is 60% hereditary is 40% so wonderful model system to study the cancer where genes genetics is also involved environment is also involved but no one has any answer ok now this is terrible make is not good to see too much ok in 6 months times this become terrible ok if you did not get treatment then wear it off and that is the problem happened in developing country when you get you see leukocarea you can characterize if a doctor will see your normal person will quickly figure out there is something in the eye and this is the step where you understand that there is something leukocarea step you will tell I have mouse retinoblastoma mouse in lab we see that directly it ok but if it get bigger they get tumor bigger than head size now you can understand how this happen if timely there is nothing inuculation is only that a chances to having metastasis is very very high so what you should do now this is the chain if and most of the time what happened children parents not able to take to the hospital no one takes care it gets different aggressive a stage and then it is almost nothing now so what we need we need a need for the development of safe better and effective manner what you need at this time you need some drug ok whatever drug is there that is also not able to cure it has very side effect then this one of the student kamachi phd student has done this work how now we thought to break the cluster ok and now idea is that find a drug and find mode of action in science mode of action is more important drug needs patient mode of action will give you patient help and to make the another drug better ok so now simple concept break the cluster you see this is cluster forming cell ok so now when you are started giving this control cluster is there you give we have screens few compounds several compounds and then we came to this compound this break the cluster but this is not a big deal you put surf excel wheel anything it will break the cluster so this is nothing conclusion but indication is there ok now if you got what the biologist will do quickly go and do cytotoxicity ok so we did cytotoxicity and somehow we found why 79 cancer retinoblastoma cancer cell line is giving 18 micro molar IC 50 but if you got 18 micro molar in the cancerous cell it is also possible this will kill your normal cell ok if it is killing normal cell then how can this will be drug first argument you ask yourself and you confirm here now what we did we took ARP 19 it is also retinal epithelial cell and here you see IC 50 is 165 micro molar that means there is it is not killing normal cell we are looking for this we are looking a compound which has potential to kill only cancerous cell not to normal cell that is the whole point came here so now you see this is our slide anyhow there is nothing hard to tell fact this experiment we do did very late first we did because we do not have cell line we try to arrange cell line it took lot of time later we did but somehow it worked now here proteomics played the role now I told you you put surf excel also it will work ok but when surf excel concentration will go it will come again cell closure ok now when this compound this never comes close so that was one benefit now you took the give the treatment with this compound to different cell ok both now you do proteomics I have shown you how to do it ok now you do proteomics now you start comparing what happened after doing proteomics when you look the proteomics what you found cell adhesion related proteins are down regulated in e4 e4 is nothing it is my lab number room number ok so that is from it came so now all cell adhesion related protein so cell is not getting attached it is getting detached and cell adhesion protein is getting down regulated that means something is going towards the cell adhesion no one is going to believe proteomics if you are not doing antibody or some validation at least reviewer ok we all believe ok reviewer will ask you now we have done antibody experiment and we have moved forward now if it is really anti cancer all oncogen and cancer related protein should be down regulated perfectly hit ok so that means it is anti cancer now you have to talk about the mechanism what happens if all mitochondrial protein you will see that is going up regulated in the treatment so mechanism is somewhere related to mitochondria who is telling proteomics now this much indication is enough for biologist to kill the or make the project work ok that is the proteomics plate the room now you do whole analysis cluster networking all you have learned you know works of must do this is very useful when you do you saw tumor suppression protein tusk 3 is getting up regulated that is the logic no suppression protein should go up cancer related protein should go down but we found this transcription factor and caspase 14 came up so that we do not have anything ok clue about that you quickly look this compound is binding with DNA or not control take ethereum bromide and just you look it is not binding quickly next experiment TM oh TM is changing somehow no need to find quickly mechanism ok one by one one by one third you are in CCMB all are well trained here ok now third quickly look does E4 after altered cell cycle here we found a little bit very different and strange result you see this sub G1 population is getting very high with the concentration dose dependency ok that means you change the dose IC50 is 18 if you increase this dose and here you find the mechanism sub G1 population is getting very high here you see in same constant thing in 24 and 48 hour result is wonderful something confusion is there this pattern this pattern is same this pattern this pattern is changing in the S phase no answer we have ok this happens that means different dose is working at different different time point at different things ok but this gives sub G1 population is altering the circuit so what is the conclusion E4 perturbs the cell cycle and cause the cell death if this we know then next what we should do quickly do apoptosis and quickly we are showing again with the concentration increasing the concentration what you are doing you are able to go to late apoptosis more so this is working in the time and dose dependent manner that is we come you cannot conclude anything more than this now E4 cause cell death by apoptosis now you go another experiment DNA fragmentation run the gel quickly do it or do tunnel as a little bit fancy and you can see that DNA fragmentation is also going up by higher concentration so we are trying to find the mode of action mode of mechanism and question answer is here E4 cause apoptosis by DNA fragmentation dose and time dependent manner even not binding with the that so now detection of intracellular ROS we have some mitochondrial thing we have saw ROS protein so with this we found that this is also making sense and mitochondrial membrane potential SA we have done with this we came to that it is something is happening we are not sure ok something is happening towards the mitochondrial cell so compound E4 induce ROS generation mechanism and leads to mitochondrial membrane depolarization this indicates that ok so what proteomics told mitochondria ok result came by other biological technique same or different that is we need so with this now no one is going to believe you cell culture and only publication ok if you want to move mouse experiment you have to do so this is genograph mouse you take the cell line inject in the mouse make the tumor leave a month time to stabilize inject the compound and if it reduce supervisor will move next stage and a student will go happy another year for move ok so this happens what is the most important thing if you give take control because after 1200 ethical doesn't permit you to keep the tumor size and if you give treatment it is completely demolished this results gives very nice even I like this is first experiment you see because experiment you have to plan very well give expert injection only on Monday a student will not feel bad Monday come inject nicely wait for next Monday and you see the once how dose dependency go so this proves clear that don't give too much pressure then they will not think ok so this gives clear that how things are working if it is really working what is the next stage again use your proteomics now we are sitting here we have this tools technique knowledge we took the tumor from both after treatment that do full proteomics do histological staining quickly so where is the apoptosis is happening and histology of tumor thus you can see treatment with just this and now I told you one word side effect there should not be a side effect ok so now when histology of liver we do we don't see any change only compound when you inject when tumor form when tumor form any compound injected we don't see in histology any change and no mouse has died that is the biggest with this with this you see similar thing in spleen you saw similar thing in lung heart and kidney that gives the now here 2 and half years after hard work of system and paperwork finally we got knockout mouse retinoblastoma where gene is knockout RB minus P53 minus so mouse will get automatically retinoblastoma here we have started treating and we are getting potential very good result and I will show you most probably next time whenever we meet with this I showed you how things will go similar concept we have used for leukemia very nice IC 50 1 1 minute I will show you the concept is repeating that you got late apoptosis what is the interesting thing here got when you injected this is another compound but similar line in 5 minute it reach after injection to brain and if you leave for 45 days this compound get from body out also so this is another compound working for leukemia this is very nice third is melanoma all you know what I saw how to come the mechanism this is the last here there is no cluster if it is working there is no cluster and when cluster is there still compound is working you find mode of action and mechanism which signaling pathway very simple all the known target drug you take make the slide and when you find this is the target and then you do all your experiment and you will find by antibody this is happening ok I told you in biology antibody is very important now you see BCL 2 is the downregulated PAKT is downregulated and if this is really apoptosis all cast pages should be downregulated or upregulated upregulated biologist like this only mass spec is not enough you need mass spec and parallel this when you reach this by this anyone can tell there is the apoptosis cast pages up there is something is going if you have this data you make your models how mechanism is working when you go to this mechanism you saw we saw cast page 8, 3, 7 poly adipure apogylation cast page 9, FF, BCL 2 backs so this is the mode of that and mostly it is going towards the AKT signaling pathway ok now use your knowledge here experience is experience ok then you see it is AKT, FOXO 1, BCL 2, P53 backs apoptosis this is the chain it is coming downregulated and upregulated so finally we are coming it is towards the AKT signaling pathway with this thank you all of you your patience and for time. From today's lecture by Dr. Suman Thakur I hope you got a complete image understanding about why getting a single biomarker for cancer is tough why many biomarkers have not been able to reach to the transitional work so far in the clinics Dr. Suman Thakur showed you how multiple cancer cell lines having same protein but different amount this is reason a labeling technique like Syrac or eye track can be helpful. He also showed a complete overview how cell biology can be used to get the drug screening strategy and unable model experiment can help in validation. I hope you are gathering different facts from different scientists clinicians and trying to understand that what are the latest advancement in the field and also what are the gaps from the clinical as well as its successful translation in which way by integration of cell biology various type of omic based technologies and clinical strategies together only we can make some success in this area. The next lecture is by Dr. David Fanu who will talk about predictive analysis thank you.


If there was one animal that exists that tells us in science what is possible in almost a magical world of not aging or that basically having any cell in your body be able to completely reform into the identical you that was before and the memories that you had prior, would you guess what that animal was? It's a flatworm. Imagine that a flatworm is everything in the most ideal world of what is thinkable in science if we could figure everything out. I talk about this with Dr. Michael Levin at Tufts University. He got all of his undergrad training as PhD at Harvard Medical School where we discussed not only what regenerative medicine is but the bioelectric network and how cells that escape the bioelectric network can actually turn into cancer. Check it out in this extremely mind-blowing episode of Target Cancer Podcast with Dr. Michael Levin.


My name is Toya today Taylor, and I'm the executive director of we app we act presenting perform the speak with purpose Program, which is the only public speaking class offered in Seattle public schools at many of your schools And we're so proud of that. I'm here with my team today. This is miss Sophie. Hey, miss Crystal And today we are working on a free ride So some of you know what a free ride is but some of you're like what in the world is that is it a free Right that you give away to somebody or what? This is an opportunity, and we love doing this and speak with purpose for you to freely express yourself It's all about letting go of judgment Allowing yourself to really just say what it is that you are feeling in the moment Nothing more nothing less. It doesn't matter how old you are how young you are We do this with students as young as fourth grade all the way up to 12th And you can't imagine the amazing pieces that people come up with when you allow yourself just to be free All right, so I'll say this for me free writing was really important when I was probably like in ninth grade Ninth grade for me was hard you guys. It was a time that I felt like nobody. I mean no adult Really listen to my voice. I felt really alone and like I Had things to say I just didn't know how to say them. Have you ever felt that way? Yeah, middle school elementary Yeah, I think at some point it was even elementary through high school Until I learned I found this this thing I could do called free writing And this became my voice it became my opportunity to talk about whatever was on my mind And I began to talk about my community my life how I felt about school And how I saw myself in the future and so I say to you if I can do this Anybody can do this and you should and we will today. All right, so first thing about free writing is I'm gonna show you How we get started first of all you want a journal If you don't have a journal snow sweat, it's not a big deal get something to write in or on so a piece of paper What else could you use? Use your phone All right, you just want something you feel comfortable writing in for me. It's a journal for today. All right We're gonna give you ten seconds to go and get Something to write in you ready. I'm gonna break it down for you a pen or pencil either one Thanks, miss. Oh, I got you. Thank you. So we're gonna break it down for you. Go get it ten Here we go nine where you at where you at eight go get a pen or a pencil seven What what six five five four? Three are here. You come to find a place so you can write one one one one One all right cool. Um now that you have something to write with Or in Show you how we do our free writes Here's a free write that I did the other day You're going to put your name in the left hand corner. So open up to any page. That's a free page write your name in the top left hand corner and Then right across on the opposite side. You're gonna put today's date whatever that is Okay Today is 424 in the studio correct and then whatever we choose as a topic or two words That's what you're gonna put in the middle and then you always want to find some place to put the exact exact time that we start and We'll tell you what time to start and you'll put the time there then All right, so put your name left hand corner Date right hand corner will tell you what the topic is today and then you'll also put the time we start Cool now.  It can be under a table on a table on the floor left up to you Next when we say pen the paper once that timer starts you do not you just write Don't stop writing. Don't think just do allow your mind to get comfortable with writing If you get to a place where you're writing and you're like, I don't know what else to write write it I don't know what else to write. This is crazy. Why am I writing? Maybe the sky will show something open up to me and I will see and be free and you'll just start let your mind Get there. Okay. The other thing is most importantly intercoach It's important that you find your voice because I can't even I can't even tell you how many songs I've Tried to write or poems and didn't finish because I thought nobody wanted to hear them or I didn't know what else to write or I just Gave up on myself That's an inner critic. That's what we don't do We want to have that inner coach that says I can speak about anything. I want the world is open to me I believe in my words somebody else needs to hear this But I have to be the one to write it or I'm just writing it for myself and that's all that matters I can do this. Okay intercoach Dispel the inner critic All right, so I think that I miss anything you guys I Think we're almost ready Okay, get your journal whatever you're writing with I want you to now go find your sweet spot To be on the floor I might just stay here because I'm the oldest of everybody at the table, right? So I think I'm a I think this feels good to me. This will be my sweet spot for today. I love it miss Sophie Oh, I like that. That's nice miss crystal So let's pick two words that we're going to free write today. Um, I'm thinking heart What are you guys thinking? I like it, too Now remember the beautiful thing about a free write is be free Use your imagination. Heart doesn't have to mean the traditional definition of the heart bumping in your are In your chest it can be a planet call heart a food call heart How could we use friends? You can think about the stuffed animals you have as friends Your stuff You're even like a pet named friend a pet named friend Friend could be the name of a school you go to the name of a song That was made on planet heart Whatever you want. That's the beauty of a free write is you choose What the words mean to you and explore it be open? Okay, so we're gonna get our journals or whatever you have to write and I want you to write in the middle of that page put heart friends Your name in the left hand corner remember and the time in the right in the time that we start We're in the studio and the time is 1135 okay We're gonna play a little music find your sweet spot. Oh, I saw nice. This is gonna be my sweet spot. Here we go Wow three minutes goes by really really fast So write your last word. I see miss crystal is still writing her last sentence. Take your time and put the time that we stopped and I'm going to Share my free right. Is that okay? All right, and I want you to remember sometimes people are like, I don't want to share you don't have to But if you want to you can share with a friend a sibling a parent Or just read it out loud to yourself So you feel really good about or you just hear the words you feel good about it And know it's for you. All right, so this is my version of heart and friends Heart feel the room flowing from speakers like trains in Brooklyn pouring out like Nina Simone or Miles Davis The song she played was friends a melody a beat that reminded me of home Reminded me of my grandmother's sweet potato pie. I cried the tears came from my eyes as I Heard her pour into the room slow like sweet molasses like sweet Magnolia flowers. I Thought about friends some old some new they all crossed my mind And in that moment I was lost in time So I appreciate it. Thank you We won't go into your free rights, but what were your free rights about? Really quickly Writing about her little brother having a friend named heart in elementary school Nice So miss Sophie used the traditional sense of Expressing your heart and having good friends in your life. There's no wrong or right in a free right So remember find your sweet spot once you start writing. Don't stop. Just allow it to flow Okay, give yourself a time. I would say three minutes to five minutes more importantly remember Always embrace that inner coach that allows you just to be yourself and express yourself to the fullest and Dispel that inner critic that tells us what we can't do because we can do it all All right, so if you want to share some of your free rights with us We would love to hear them We're gonna put up our website at the end of the video and you can share them right on our site And we would love to hear some of your free rights. Yeah Cool My name is Toya today Taylor. I'm the executive director of we out. This is Enjoy life be true to yourself speak with purpose and we'll see you in the next video. Bye


Would you please tell us Your testimony and how did you receive your healing and what were the things you were struggling with in the first place? Well, I was struggling for a few years with quite a few things COPD asthma and then the affliction of vertigo and I went through many many many doctor visits and Was prescribed lots of medicine that I came to accept the fact that I was Gonna live like this for the rest of my life and deal with it now how did I get my? Healing quite by accident to be honest with you my oldest daughter and Wanted to come to the Bob Larson conference and she lives in in another state And came to stay with me to get ready for it and said mom You want to go and I go no she said well, you know it ain't gonna hurt No, I don't want to go so the day before later in the afternoon. She come to me and said mama Don't you want to go see and meet the people that your granddaughter Haley Has been involved with And I was sold lock stock and barrel I'm gonna go meet these people I'm gonna see what my granddaughter is doing So I went And my youngest daughter showed up with my other granddaughters and we listened to the service and it was time for Bob to perform his deliverance and They called the people up to the front and I Never Intended to go there whatsoever, but my daughter ran As fast as she could up there And being her mom.  She's gonna need my help I'm going up there with her. I'm gonna be standing there with her. I'm gonna catch her as she falls and We're standing there She's crying and crying and I noticed behind me a young man of God from this church Come in behind checking And then something happened to me that I still have no idea about And I quit trying to cast gas question or anything else a Fog came across My head just fog. I Looked at my daughter and said I Gotta go I got to leave her right now And I started to leave and my daughter grabbed me by my arms and said no mama Mama stay Please stay And I was fighting her to get away and By that time the young man had approached me and touched me and My spirit was in me and I could see and I could hear But the words coming out of my mouth were not mine And I was hollering and yelling that this young man was not gonna touch me. No He's not gonna touch me And of course I was raising quite a wreckus and so they kind of asked me to leave And get out of there So that was kind and they took me and set me down On the side and this young man kneeled down before me and touched me on my knees and He was praying and praying and praying and I couldn't understand a thing. He said But he was praying And I felt something that I have never felt before in my whole entire 65 years a warmth from my feet Started to engulf my body and the fog lifted Come on, let's put our hands together for Jesus Christ Awesome, what happened afterwards? well, I Looked at that young man and said who are you and He said I am a man of God And then he escorted me out to a foyer and he stayed with me and he prayed with me And they prayed with my daughters and my granddaughters and at that day Not asking or expecting anything my family was healed not just me my family Come on. Come on. This is awesome Wow, praise God And can you share with us? How is your health now and the things that you were struggling with? How did you get healed? I walked out of the Bob Larson conference. I Take a deep breath without coffin. I Could walk with no pain. I mean we were getting ready to buy me a wheelie cart to I get around I Walk I went with my granddaughters yesterday to the mall for five hours and walked Yes Well So now the medication that you're holding can you please share? What was it for? The one in the green is for COPD. It's called QVAR That I had to take once a day twice a day This one here was every four hours or More if I needed it for asthma This one here is Brand new completely full and I've had it and never had to open it This one here. I stopped using it that day and haven't touched it again Put your hands together for God Wow, praise God So can you please share? How did those sicknesses affected your daily life? What you couldn't do that you now can't do I couldn't do nothing I mean, I'm sorry. I mean to put it My brother who lives with us had to vacuum my floors and sweep and scrub them Because I couldn't I didn't have enough breath To do it I could not do it To walk to do anything at any minute I could not do it absolutely no way and now I am up and I've already got my bathroom redone and Restained and painted and done and working on my kitchen cabinets next and Going to the mall with my granddaughters. Wow God is good a man Patricia, can you please share one thing that you shared with me before the service about the ear problem that you had please and what happened? that was an attack I had at my grandson's graduation party and it hit me and it floored me and It is behind your inner ear is a little bone and it helps your balance and Mine's gone and so if I get too hot Turn my head this way and I'm doing it this way because I could not do that Had I done that before it would have dropped me on the floor and I I Can do it Wow Now she mentioned to me that she always had to have another person in a car with her because she could not turn Her head to look if there is another car driving this way or that way and Right now. Do you have to have a person in your car to turn your head? Not at all Not at all. Amen and praise the Lord and I am so grateful Put your hands together for God. What a powerful powerful God Now with all these amazing things that happen to you Would you please share a word of advice to people who might find themselves in a similar situation With a family problem or sicknesses or in need of deliverance. What would you suggest? If anybody has any affliction whatsoever pain, this is not from our father Give it to the Lord Completely give it to the Lord and know in your heart and feel it and pray Because he is the healer of all healers


Tropical Storm Karen is poised to make landfall sometime this weekend on the Gulf Coast of the United States. Tropical Storm warnings and hurricane watches have been issued for the area. We recently spoke with Jeff Sanford, the Ready Navy program manager about what it takes to be ready in the event of a natural disaster or emergency. Except for, everyone knows about being informed, making a plan, right, having a kit. We're adding that fourth one about staying informed, it's not just being informed, but always stay informed. For more information, ideas for items to include in your preparedness kit, and for tips to keep your family safe, log on to ready.navy.mil. From the Defense Media Activity, I'm Petty Officer Tony Rosa.


The seven most beautiful cities in Italy for teaching English abroad. Italy is one of the most popular destinations for teaching English in Europe, and it's not hard to see why. With so many amazing cities to choose from you will be spoiled for choice in this Mediterranean paradise. Tips for teaching English in Italy Because of strict EU laws and regulations, it can often be difficult for non-EU citizens to secure a visa and work legally in Italy. However, if you are dreaming of living and working in Italy, don't worry. Check out our detailed guide on how to legally teaching English in Italy for people outside of the European Union. Here are the seven most beautiful cities in Italy for teaching English abroad. Teaching English in Bologna The capital of Italy's Emilia-Romagna region, Bologna is a premier destination with a lot of exciting attractions. Not only is Bologna stunningly beautiful, it's also home to the oldest university in Europe. Moreover, Bologna is famous for its very own Leaning Tower, it's not just in Pisa. Try your way through Bologna's local cuisine at the mini-tratoree and mingle with the locals to learn Italian. Teaching English in Florence One of the most visited cities in Italy, Florence has inspired countless artists and writers due to its unique architecture and romantic vibe. The city is most famous for its Duomo in Palazzo Vecchio, two true Italian gems. While the cityscape of Florence is utterly beautiful to look at from the outside, don't forget to step into its many historic buildings to see works from masters such as Michelangelo, Leonardo da Vinci, Raphael, Titian, Caravaggio, and more. Teaching English in Milan Located in the northern Lombardy region of Italy, Milan is considered an international hub for fashion and design. Besides that, Milan also houses the Italian Stock Exchange, making it an important business and financial hub in the country and the whole of Europe. The city is also known for its mini-high-end shops and restaurants and the extravagant lifestyle of many of its citizens. The Duomo di Milano is the city's most important landmark and a perfect example of Gothic architecture. Art fans should also visit the Santa Maria del Grazie Convent, which houses the world-famous mural The Last Supper by Leonardo da Vinci. Teaching English in Naples While Naples often has a rather chaotic reputation, it's a city that surely shouldn't be missed when in Italy. The city's world-famous cuisine is worth the visit alone with its pizza, pasta and seafood dishes of extraordinary quality. Napoli is also considered one of Italy's most important art and archaeological locations and offers top-notch museums to locals and visitors alike. Plus, you're never far from the beach. Teaching English in Rome The capital and largest city in the country is among the most visited cities in the entire world. With a history of more than 2,500 years, there's undoubtedly a lot to see and do in Rome. Whether you're interested in the archaeological sites and art museums or want to check out as many churches and ruins as possible, Rome has got you covered. Don't forget to stop by the Vatican during your visit to see world-famous sites such as St. Peter's Square and Basilica and the Vatican Museums. It's true, all roads lead to Rome. Teaching English in Venice Surely, there is no question why Venice is considered one of the most beautiful cities in Italy, or even the world. Just take a look at the unique city outline built entirely on canals. All around Venice and you won't have any traffic but all you hear is the sound of water from underneath its buildings. Adding to that, you've got unique architecture, beautiful churches, and the eye-catching St. Mark's Square. Teaching English in Verona True Shakespeare fans will not want to miss a chance to visit the city where William Shakespeare said Romeo and Juliet. Besides its importance in world-class literature, Verona also boasts fascinating medieval castles, piazzas, and historic churches, along with ancient Roman ruins that are incredibly well-preserved. Verona is a deal for romantics and those looking for history.  Speak with an ITTT advisor today to put together your personal plan for teaching English abroad. Send us an email or call us toll-free at 1-800-490-0531 to speak with an ITTT advisor today.


Good evening and welcome to this distinguished lecture, which is co-organized by the Food Studies Center and the Agrarian Change and Development Research Cluster, both here at SOAS. The SOAS Food Studies Center is committed to the study of the political, economic, and cultural dimensions of food, and more specifically to the dynamic interaction between these dimensions, whether in the past, present, or future, from production through to consumption, not only in Africa, Asia, and the Middle East, but also in the rest of the world. The Center fosters the teaching of food-related courses at SOAS, facilitates supervision of research theses, and builds networks between scholars with an interest in food at SOAS and beyond. More than 40 members of SOAS staff belong to the Center, as do more than 20 SOAS research students, and more than 100 students on talk courses at the school and alumni of these courses. Its several hundred associate members include students and academics from other universities, policy makers, activists, journalists, and makers and vendors of food. Among its most important public functions are the weekly SOAS food forum and distinguished lectures such as tonight's. Should anyone wish to join the Center as an associate member, which is free of charge, and to be placed on our email list, please send an email to soasfoodstudies at soas.ac.uk. The agrarian change and development research, the agrarian change and development research cluster encourages investigation of the social relations and dynamics of production, property, and power in agrarian formations, and their processes of change, both historical and contemporary. It works within a comparative, within a broad interdisciplinary framework, informed by theory, and generates rigorous comparative analysis and scholarly debate on agrarian structures and change, past and present, in different parts of the world. The cluster functions as a reading group and a sounding board for the ideas and interests of its members who are drawn from the departments of development studies and economics at SOAS. The cluster meets several times each term and members, including staff and research students, discuss early drafts of papers, pieces they've recently published, or texts that have attracted their attention. The cluster works in tandem with the agrarian change seminar, convened it so as by the editors of the Journal of Agrarian Change, which serves as a more formal public venue for the presentation of research papers. So on behalf of my colleagues in the Food Studies Center and the Agrarian Change and Development Research Cluster, I wish to thank the Centers and Programs Office here at SOAS, especially Nena and Jane, for all of their hard work in organizing the event tonight. We also wish to acknowledge the support provided by the Journal of Agrarian Change, as well as by Gastronomica, the Journal of Critical Food Studies. For those of you who are not yet aware of it, this evening's speaker will be back tomorrow from one o'clock to three o'clock in the Khalili Lecture Theater to respond to questions about his work, which have been submitted in advance by SOAS students. Should you be interested, you are most welcome to attend. So let me now introduce our speaker. James Scott is the Sterling Professor of Political Science and Professor of Anthropology at Yale University in New Haven, Connecticut. His research concerns political economy, comparative agrarian societies, theories of hegemony and resistance, peasant politics, revolution, Southeast Asia, theories of class relations, and anarchism. He studied political science at Williams College, where he took his BA, and then after time spent studying economics at Rangoon University in Burma, and political science at the Institut des Sciences Politiques in Paris, he took an MA and then a PhD in political science at Yale. He first lectured at Wesleyan University in Connecticut. He then spent time at the University of Wisconsin in Madison before returning to Yale, where he became the founding director of the program in agrarian studies. He is author and editor of a great many books and scholarly articles. His major publications include The Moral Economy of the Peasant, published in 1976, Weapons of the Week, Everyday Forms of Peasant Resistance, published in 1980, Domination and the Arts of Resistance, published in 1985, Seeing Like a State, How Certain Schemes to Improve the Human Condition Have Failed in 1998, The Art of Not Being Governed, An Anarchist History of Upland Southeast Asia, published in 2008, and most recently, Two Cheers for Anarchism, published in 2012. Sorry, another book decoding the subaltern, Ideology, Disguise and Resistance in Agrarian Politics, also 2012. He's held numerous scholarships and fellowships. He has served as president of the Association of Asian Studies from 1997 to 1998, and he was named a Fellow of the American Academy of Arts and Sciences in 1993. According to his vita, he is also a member of the Connecticut Sheep Breeders Association. Yes, Jim, I was paying attention. Those who know Jim have long valued not only his scholarship, which is at once meticulous and engaging, empirically grounded and theoretically stimulating, but also his generosity in sharing his time and ideas, whether as a teacher, as a colleague, or as a mentor of young scholars who for decades have nestled into the program in agrarian studies, often to write their first books in this intellectually fertile niche. So it gives me great pleasure to introduce Professor James Scott. Thank you, I'm delighted to be here, and also to be at an event sponsored by the SOAS Food and Agriculture program. I think of it like many others as the children of agrarian studies at Yale now in its 22nd year, and most of these children, I should add, have far outstripped their parents, right, in terms of their reach and their intellectual influence, and I think it's what every parent hopes for from their children. The talk I'm giving tonight is from a yet uncompleted book that was inspired by my realization that everything that I thought I knew about early states and the domestication of grain and sedenturization was basically wrong, and this is my effort to provide an account that's more satisfactory for myself and hopefully for others. The other potential title of this talk could be the late Neolithic multi-species resettlement camp. Little by little, I've tried to develop an account of how we get agrarian societies in the first place. After all, Homo sapiens has been around for about 200,000 years, and only 60,000 maximum of those years out of Africa. The first agrarian states by then just a mere smudge on the map and a rounding error in the world's population appear at a generous estimate roughly 6,500 years ago. They represent just a shade over 3% of our history as a species, and I was somewhat surprised to learn evidence of domesticated food plants and livestock occurs several thousand years before anything that we could call an agrarian state pops into archeological view, and it's this gap of 4,000 years that in part preoccupies me. How might one tell the story of how we as a species, having spent virtually our whole span on planet Earth as hunters and gatherers, were assembled in great clumps, growing grain, tending livestock, and governed by the political units we call states and empires? One option and the one I choose to exploit here is to examine the preconditions that made it even conceivable that we should gather or be gathered in substantial numbers in a single place and not immediately starve. These preconditions are, I believe, the result of three world-altering domestications, fire plants and animals. Each of these three domestications brought us closer together, reshaped the natural world, and in turn reshaped us as well. In domesticating much of the natural world, we in turn domesticated ourselves. Only in the context of these domestications did we become suitable raw material for the critical concentrated mass of foodstuffs and manpower that state formation requires. The state is, as we shall see, both the beneficiary and the forcing house of these domestications. What fire meant for hominids and for the rest of the natural world is thrown into sharp relief by a cave excavation in South Africa. At the deepest and therefore oldest strata of this cave, there are no carbon deposits and hence no fire. Here, one finds full skeletal remains of large cats and fragmentary bone shards bearing tooth marks of many fauna among which Homo erectus. At a higher and later stratum, one finds carbon deposits signifying fire. Here, there are full skeletal remains of Homo erectus and fragmentary bone shards of various mammals, reptiles, and birds, among which a few gnawed bones of large cats. The change in cave ownership and the reversal in who was apparently eating whom testify eloquently to the power of fire for the species that first learned to use it. At the very least, fire provided warmth, light, and relative safety, as well as a precursor to the domus or the hearth. The case for the domestication of fire being the decisive great transformation in the fortunes of hominids is, I think, convincing. It's been mankind's oldest and greatest tool for reshaping the natural world. Our ancestors could not have failed to notice how natural wildfires transformed the landscape, how they cleared older vegetation and encouraged a host of quick colonizing grasses and shrubs. Many of them bearing desired fruits, berries, and nuts. They could also not have failed to notice that a fire drove fleeing game from its path. Exposed hidden burrows and nests, and more important, later stimulated the brows and mushrooms that attracted game. In native North America, this would mean elk, beaver, hare, porcupine, turkey, quail, rupt grass, for example. Native Americans were harvesting, if you like, a suite of wild game that they had attracted by their use of fire. Once in control of fire, early mankind, until today, hunters and gatherers and swidden farmers, have used fire to transform the landscape more to their liking. They encouraged the flora and fauna they desired and discouraged those they didn't. Until roughly 100 years ago when fire was declared, at least in America, an enemy on public lands, it was scarcely, there was scarcely any part of the globe, including the rainforests of Amazonia that had not been remade by human, that is to say anthropogenic fire, and hardly a landscape that was not fire adapted. Here I just wanted to show very briefly, this is a print of the hunting of kangaroo by Australian aborigines, early in Australia's European history of Australia. This is a burned swidden. This is, of course, a swidden fire. And finally, this is a landscape that shows you the swiddens that are recently cleared and fallow for different periods of time. We best understand this massive human intervention in the natural world, I believe, as low intensity farming and animal husbandry. It is a kind of horticulture done over millennia over a long, long period of time that in the long, although by imperceptible degrees, completely transforms the landscape. For our purposes, however, the wielding of fire has the effect of assembling, gathering, and concentrating selected resources closer at hand. Fire acts as a powerful magnet, bringing a suite of desirable flora and fauna closer to the hearth. We're a very long way, of course, from the grain field or the livestock corral here, but on a very long view, one can see the centripetal force of fire accelerating the reorganization of the natural world for the convenience of Homo sapiens. Fire served then as the plow and harness of biotic domestication for about 500 millennia before what we know as the Neolithic revolutions of farming and animal husbandry proper. Rather than roaming far and wide for naturally occurring food, mankind has now so rearranged the world, so as to surround herself and put her needs closer to hand. Concentrating foodstuffs and game reduces in proportion the necessary radius of hunting and gathering. As the carrying capacity of a given landscape grows, the population potentially becomes slightly more dense and potentially more settled. Fire powerfully concentrates people in yet another way, cooking. It's virtually impossible to exaggerate the importance of cooking in human evolution. The application of fire to raw food externalizes the digestive process. It gelatinizes starch and denatrous protein. The chemical disassembly of raw food, which in chimpanzees requires a gut roughly three times the size of our own, allows Homo sapiens to eat far less food and expend far fewer calories extracting nutrition from it. The effects are enormous. It allowed early man to gather and eat a far wider range of foods than before. Plants with thorns, thick skins and bark could be open, peeled and detoxified by cooking hard seeds and fibrous foods that would not have repaid the caloric costs of digesting them became palatable. We extract, if we eat raw grains, for example, the diet of most Homo sapiens, if we eat wild grains, excuse me, domesticated grains raw without cooking, we can extract only 10 to 15% of their nutritional value. They need to be cooked in order to provide us with the nutrition we require. Even before the advent of cooking, Homo sapiens was a broad spectrum omnivore, pounding and mashing raw meat and plants, but with fire, the range of foods that could be digested rose exponentially. As testimony to that range, an archeological site in the Rift Valley dated 23,000 years ago, gives evidence of a diet spanning four food webs, water, woodland, grassland and arid zones, encompassing at least 20 large and small animals, 16 families of birds and 140 kinds of fruits, nuts, seeds and pulses, as well as plants for medicinal and craft purposes. Fire for cooking was at least as important as fire as landscape architect for the concentration of population. The latter placed more desirable foods within easier reach while the former rendered a whole range of hitherto indigestible foods now both nutritious and palatable. The radius of a meal, you could say, was greatly reduced. Not only that, but softer cooked foods as a form of external premastication allowed earlier weaning and was almost certainly responsible as long with sedentism for the enormous demographic expansion of agricultural people's vis-a-vis hunters and gatherers. The genetic and physiological effects of at least a half a million years of cooking have been enormous. Compared to our primate cousins, as I mentioned, our gut is well less than half their size, our teeth are far smaller, and we spend far fewer calories chewing and digesting. The gains in nutritional efficiency, it is claimed, largely account for the fact that our brains are three times the size one would expect by comparison with other mammals. In the archeological record, the surge in brain size, I read, coincides with hearts and the remains of meals. Morphological changes of this magnitude have been known to occur in other animals in as little as 20,000 years following a dramatic shift in diet and ecological niche. Fire, one might say, is our trump card as a species, a monopoly that largely accounts for our reproductive success as the world's most successful invasive. Much as trees, plants, and fungi, we too are fire-adapted species, pyrophytes. We have adopted our habits, our diet, our body, to the characteristics of fire, and having done so, we are chained, as it were, to its care and feeding. If the litmus test for a domesticated plant or animal is that it cannot propagate itself without our assistance, then by the same token, we have adapted so massively to fire that our species would have no future without it. One small but telling piece of evidence is that raw foodists who insist on cooking nothing invariably lose weight. Amir, that's a fact that satisfies me. Amir, 10,000 years ago, all Homo sapiens, roughly 4 million of them, some people judge, were all hunters and gatherers. Between 60 and 80% of our diet came from plant sources supplemented by nutritionally valuable high protein foods from hunting, fishing, and shellfish gathering. Not long afterwards, there's evidence for the domestication of wheat, barley, millet, maize, lentils, potatoes, and peas from a relatively few centers where their wild ancestors are found in the Fertile Crescent, Southwest China, the Andes, Central America, and Ethiopia, and here are just for purposes of illustration. This is emmer wheat, the sort of earliest wheat found along with einkorn in the Middle East. Here is a demonstration of the difference between wild emmer wheat on your left, which shatters easily and therefore does not wait for the harvester and has a very brittle rachis and a domesticated emmer, which has a less brittle rachis, which means it doesn't shatter and can be brought back and thrashed by Homo sapiens. You can see the differences also in the size of the grain. The little box in the middle is the, where the earliest wheats are found and then the rest of the red represents the major wheat growing areas subsequently. And finally, this is an American quarter for size of comparison, of the comparison to the Teosinte plant, which is the wild ancestor of maize, and then the sort of modern maize plant and you get some idea for both the enormous increase in the numbers of seeds and their size over time and the changes in the plant. We'll talk later. This is an example of flood retreat agriculture, which is the easiest possible agriculture to practice historically, and this is sorghum growing there. These domestications that I mentioned did not precipitate the rise of agrarian civilizations. In fact, as I mentioned, it would take at least four millennia before these grains would become the staples of the first agrarian kingdoms. This long interim is crucial. Not only did the domestication of plant foods long predate agrarian states, but all of the 12 to 15 crops that today dominate world commerce were domesticated by the sophisticated applied botanists, also known as our Neolithic ancestors. Historical man has added virtually nothing to this achievement. What is perhaps most astounding about our diets in the subsequent 10,000 years is how narrow they have become. To the point where they are dominated, I think 60% of the total calories consumed by humans today are dominated by the three major grains of rice, maize, and wheat. As someone said, we've all become parrots. The question of why early farming was taken up is intrinsically more interesting than how it was invented. Mankind has been cultivating and changing the landscape for a very long time, and the act of domestication in terms of dropping a seed in a small hole in the ground, the more you take that as the decisive rubicon of domestication, the more trivial it seems when looked at the transformation of landscape and the cultivation and weeding and care of wild plants over time and their transplantation. It's not obvious why hunters and gatherers provided with the bounty of the natural world and made more accessible by fire would choose voluntarily to move toward farming with its far more demanding regimen of labor. Even in the 20th century, Marshall Solombs has reminded us that foragers and hunters who are in marginal zones that are disfavored by agriculturalists that they could still be described as the original affluent society. They typically spend less than half of their waking hours in what we might call work, and the rest of the time in repose and in what an anthropologist might call cultural elaboration. The dominant though not entirely convincing explanation is that farming was adopted in those settings where population pressure on natural sources had increased to the point where it became necessary to extract more calories from the same territory. A theory famously laid out by Esther Bosrop nearly a half century ago. Three pieces of fragmentary evidence appear to bear out this reasoning, which I think is still quite disputable. First, in the two millennia or so before farming was established, foraging populations all over the world seemed to have grown substantially, and this is often calculated to be a result of the end of the cold snap known as the Younger Dryas, which ended in 9,600 years ago, after which conditions became warmer and wetter and very quickly and population expanded. This mini-population boom was accompanied by more intensive gathering of plant foods, nuts, grains, tubers, at the expense of meat and large game, which was presumably becoming more scarce. Starchy foods were becoming a staple before they were cultivated. A second straw in the wind is the fact that a good many sites of early agriculture were settings where crops could be sown with a minimum of labor. The most striking case is what is known as flood retreat agriculture, or Décrue in the French version. Broadcasting seed on soft and fertile silt left on a river bank by the receding annual floods. This, of course, was the Nile Valley practice from the beginning. Finally, there's abundant evidence from many sites that wherever there has been a dramatic drop in population because of epidemics, war, or famine, the remaining population typically shifts toward less intensive labor-saving means of subsistence now that the pressure on resources has plummeted. Thus, many Native American groups moved back to hunting and gathering after catastrophic epidemics brought by the Europeans. Similarly, following the black plague of the 14th century, areas once intensively cultivated were abandoned in favor of less intensive slash and burn swidden cultivation. The driving principle of Beausrop's account is the economic principle of the least effort for a given output. A principle documented in brilliant detail by Cheyanov in his study of the Russian small-holding peasants. Precisely because intensive farming generally involves higher labor costs per unit output, it's avoided so long as less onerous forms of subsistence are available. It's taken up reluctantly by hard-pressed foragers who have reached, who know how to plant crops, but have reached the carrying capacity of their environment. On this account, the adoption of farming calls into question any simple narrative of human progression from hunting and foraging to swiddening and then to agriculture proper. Agriculture almost certainly entailed a large increase in drudgery and, as we shall see, declines in health and life expectancy. Looking backward, farming seems to be man's first major step toward civilization. It cannot, however, have looked that way to those who first embarked on it. I think I have, yes, just as an attract, this is a Rosa Bonner painting of oxen cultivation, the most labor-intensive when you take into account the care and feeding and fodder required for the oxen as well as the plows as well as the work of turning over thick loam soils. I think the standard narrative historically and the narrative that I was a captive of, if you like, was that the domestication of grains and irrigation provided the first premise for widespread sedentarization and that sedentarization provided the possibility for larger concentrations of population, the division of labor, towns, and eventually the state. Now, this is the sort of narrative that I think every schoolgirl and schoolboy kind of learn implicitly or explicitly. And it beggars the imagination that through a 12-step detoxification program that you would need to get this out of people's brains, but I hope in this little book to make a start on that detox regimen. Settentarization does not require domesticated field crops. State formation does. Settentarization begins in the southern Mesopotamian wetlands. These are areas that, although now they are dry and arid, were flooded 9,000 years ago and water came all the way to Nasseria and to the doorstep of ore and iridu and uruk. The water level was much higher and civilization began, or sedentarization began. Let's put it that way. Settentarization began in these wetlands on what some people call turtlebacks, known in New Orleans and Louisiana as Shedye, these small little deposits of sand and silt that build up above the flood level, often only by a meter or so. These areas were unbelievably rich in terms of being at the seam of saltwater, marine environments, and freshwater environments, each of which had all series of subsistence resources that were different from one another. And if you lived in the tidal zone, you could say that the saline ecotone moved past you every day and then receded as the tide receded and the freshwater intruded again. The same was true on an annual basis. The area was inundated in the early part of the year and starting in May, it started to dry out, which produced an enormous amount of fodder for the sheep and goats that the people in this area had already domesticated. It was then a kind of, as some people have said, this was the Garden of Eden from which we get the sort of historical biblical story. And only later did it become an arid zone. And this history has been now, I think, documented in great detail by remote sensing, by the reading of sediments and soils. One can reconstruct both the sort of activity of foraging, of the gathering of reeds and so on from this early period. There were towns, but there were no states. There were towns of 1,000, 2,000 people that lasted for the better part of a millennium. The graves that we find are graves in which there are no differentials in grave goods, so they appear to be a relatively unstratified society. And of course these people were building with reeds and they left no monumental architecture for the archeologists to obsess over. The, I will return to the, this is the new air and the cattle. They're out of water, I'm afraid. So this is a very bad kind of slide of the area that I'm talking about in south of Basra in contemporary Iraq. It shows the marshy areas most recently and historically as well. And this is an earlier depiction of the areas that were heavily flooded historically and the alluvial fans that provided such rich soil. Area looks much like this, like this. Here you can see the marsh Arabs living in the remnant of the ecology that I'm describing. You can see the wild pigs, if you look carefully, right? In the foreground and you can see the reed buildings in the back. Here's another picture of this area. This is again, marsh Arab dwellings all built out of reeds. Many of them floating islands and some of them on these turtlebacks or chenille. You can imagine how difficult and impossible this was for the state to administer or to control. And this is a Sheik's Hall. I thought it was a mosque first, but it's a Sheik's Hall. It's a magnificent architecture all built with reeds and they have to put a new layer of floor in of reeds every year because the bottom is always being, is rotting because it's essentially a floating building on extremely wet soil. But the architecture is mind-bogglingly beautiful. After Saddam Hussein drained the marshes, this is what it looks like. I added some shellfish middens, which are very common in this area as well. This is not from the Mesopotamia, but you get some idea for the enormous depth of people who live as gatherers on the strands of marine environments that shellfish are enormously important as a part of their diet. And it was another sort of middening which archeologists are busy working. Everywhere the adoption of agriculture was protracted and halting. Domesticated cereals have been known for at least 10,000 years, but in most places it took several thousand years before they were systematically planted in tilled fields. Barley and wheat were only planted in Britain 5,000 years ago. It's not as if once shown the magic of farming, foragers and hunters rushed to embrace the advantages of agriculture. The principal reason was that a subsistence spread across several food techniques and food webs was far more stable and reliable than an exclusive reliance on planting. Early farmers might do some planting, but it was likely to be overshadowed by foraging of wild foods, collecting shellfish, trapping fish and hunting. With such a balanced portfolio, their subsistence risks were spread in a way that protected them from the failure of a single source of food. They were unlikely to give up this source of security until they were forced to. Left to their own devices, they avoided intensive agriculture until they pressed hard against the carrying capacity of their environment. When obliged to cultivate, they first opted for forms of agriculture like flood retreat agriculture that involved the highest return for the least labor. Depending on the context, they might move then to shifting cultivation or fire field cultivation with long fallow. If pressed still further, they might shorten the fallow and at last resort would take to the plow. Throughout this long transition, there was much oscillation back and forth depending on the conditions of wildlife, depending on the conditions of wild subsistence food and their availability. And whenever the pressure on land was relieved, usually because of catastrophic mortality, the survivors returned to letting fire do the work of the plow or even to hunting and foraging. The early agricultural field is a vastly more simplified and cultivated zone than the world outside it. It represents a kind of floral zoo. The purpose of the cultivated field and of the garden is precisely to eliminate most of the variables that would compete against the culture. In this artificial environment, other flora exterminated for a time by fire or the plow or opportunistic weeds pulled out by their roots or hode, birds rodents and browsers scared off or fenced out, perhaps carefully watered and fertilized, we make a nearly ideal world in which our favorites will thrive. Steadily by cuddling the plant and rigorously favoring those varieties and only those varieties that meet our needs, we often create a fully domesticated plant. Fully domesticated means simply that it is in effect our creation. It can no longer survive without our attentions. In evolutionary terms, a fully domesticated plant has become a super specialized floral basket case and its future is entirely dependent on our own. If it ceases to please us, it will be banished and almost certainly perish. We think of ourselves homo sapiens as the agent in this narrative. We domesticated the potato, we domesticated maize, rice, et cetera. But if we squint at the matter from a slightly different angle, it's we who have become domesticated. Michael Pollan puts it roughly this way in his sudden appersu while he's gardening. As he is weeding and hoeing around his tomato plants, it dawns on him that he has become the slave of the tomato. Here he is on his hands day after day, weeding, fertilizing, protecting and in general, reshaping the immediate environment to the utopian expectations of his tomato plants. Who is doing the bidding of whom becomes almost a problem in metaphysics? It's useful I think to appreciate in a larger sense how the domestication of plants as farming enmeshed us in an elaborate annual set of routines that organized our work life, our settlement patterns, our social structures and our ritual life. From field clearing and preparation by fire, plow and harrow to sowing, cultivation, weeding, to constant vigilance as the field ripens, the crop organizes much of our timetable. The harvest itself sets in train another sequence of routines. In the case of cereal crops, cutting, bundling, threshing, gleaning, separation of straw, raking, winnowing, sieving, drying, sorting, most of which has historically been coated as women's work. And by the way, if you want to know what whether the skeletal remains that you find as an archeologist were skeletal remains of women from grain civilizations, as opposed to outside of grain civilizations, all you have to do is to look at their toes because all of the women in grain civilizations spent a lot of their time on their hands and knees, rocking back and forth, grinding grain with their toes curled under, and there's a particular arthritic condition that those toes acquire over time that is characteristic of women in grain civilizations. The daily preparation of grains for consumption, pounding, grinding, fire, fire-making, cooking, and baking throughout the year sets the tempo of the domus. These meticulous, demanding, interlocked, mandatory annual and daily routines, I would argue, belong at the very center of a comprehensive account of the civilizing process. They strap us to a minutely choreographed routine of dance steps. They shape our physical bodies. They shape the architecture of the domus. They insist, as it were, on certain patterns of cooperation and coordination. Once Homo sapiens takes that fateful step into agriculture, she enters an austere monastery whose taskmaster consists mostly of the demanding genetic clockwork of the plant itself. Norbert Elias writes convincingly of the growing chains of dependency among ever denser populations that made for mutual accommodation and restraint that he calls the civilizing process. But literally thousands of years before the changes Elias describes, man was already disciplined and subordinated by the metronome of his own crops. Compare this, by the way, this single metronome of the crop to the life of hunters and gatherers who are, in a sense, judging and observing and responding to the clocks of many different forms of grain and plants and mass fruiting and so on. That is a world that, if you like, is there's not a single metronome, but there are hundreds of different metronomes that coordinate the activities of hunters and gatherers who have this wide web of subsistence resources that they avail themselves of. If fire served to concentrate more food resources within an easy reach, the transition to farming marked an accentuation of this trend. Like a powerful magnet, fire and planting shifted foodstuffs in denser and denser rings around the habitation. The planted field itself is a herbarium, adapted to the special needs of the coltogen, being nursed to maturity, and kept separate from surrounding nature at a great cost in labor. Most of this is defensive labor, if you like, to the degree that the crop is fully domesticated, the more it needs in the way of defensive work to prevent it from being overwhelmed by disease, weeds, and vermin. The shift in emphasis from pure hunting and gathering to shifting slash and burn cultivation to sedentary fixed field farming is a shift biotically from relative complexity to relative simplicity. The hunter-gatherer encourages, usually by fire, certain plants and animals he or she aims to harvest. The swidden cultivator, also using fire, encourages and plants a great variety of crops and may very well use the field as a hunting ground. In these respects, the tempo of the hunter-gatherers and swindlers is attuned to these various tempos I mentioned. From the early Neolithic domestications of grains until the advent of major grain civilizations, a very long period, people moved back and forth between these different food webs as circumstances dictated. The move toward more intensive fixed field farming was, however, a qualitative change. Subsistence production came to depend on a few staple crops that required careful tending. As food production and diet became narrower and simpler, it became by the same token more fragile. The fate of more and more people rested on the annual fate of a handful of staple crops. We had trained to live in our jealously guarded herbarium. The best way to look at domestication of animals is in the context of our concern with concentration and sedentaryism, and that is to see it through the lens of cooking. What cooking does is to allow us to eat things we otherwise could not and for less caloric expenditure. Most domesticated animals do exactly the same for us. They are our dedicated four-footed foragers, well in the case of chicken, ducks, and geese, two-footed foragers. We, in effect, send them out to forage on our behalf. They range farther than we might, and most important they eat and find, digest plants, nuts, fungi, and insects that we either cannot find or will not eat. They metabolize all of these products of nature and turn them into protein and fat, which we crave and consume. It's as if we had a large number of servant foragers scouring the environment and processing its offerings, in effect metabolizing or cooking it up and bringing it back to the hearth. In principle, the domesticated animal does no more for us than the animal brought down by a hunter. A deer provides much the same service. On the other hand, the domesticated animal, however, extend beyond the uses of the slain animal. We also make use of the domesticated animal's reproductive functions.  That milk could be preserved by turning it into such products as yogurt, butter, and cheese. In the case of poultry, the unhatched eggs could be regularly eaten. From the fur-bearing animals, especially sheep, the wool could be taken to process or weave into rugs, garments, or felt. One can skin an animal only once, but one can shear it annually. The systematic use of products of the living animal, particularly its reproductive functions, is what sets animal domestications apart from intensified hunting. And although I don't have time to make the argument here, I think I might argue that the way to understand domestication is the way in which Homo sapiens takes over and manages the reproductive functions of plants and animals that are domesticated. That is to say, to change the reproduction and control it of plants in order to produce the seeds and fruits that are desired to, in a sense, take over what had been wild animals, herd animals, and to organize their reproductive functions so as to maximize the amount of wheat or other products, or, excuse me, meat or other products that they produce. And I would argue that if we are to understand domestication as the control over reproduction, it is fruitful to extend it to the first agrarian societies in which you have property relations in the patriarchal family, in which the reproductive services of women become extremely central and their productivity in producing laborers and, if you like, domesticated animals for the patriarchal family is absolutely central as well. And it's when you get the domus and you get the statues of women in which their reproductive functions are exaggeratedly represented in the sort of neolithic statues of women that, I think, points in that direction. As with plants, the gradual incorporation of herd animals into the extended household is best, perhaps, described as a mutual domestication, recalling the domus in domestication, and selecting continually for the traits that we desired, particularly docility, we end by creating, as with domestic plants, a species that would be hard pressed to survive on its own in the wild. As a sheep breeder myself, thank you for mentioning that, Harry, for more than 20 years, I've always been personally offended when sheep are used as a synonym for cowardly crowd behavior and a lack of individuality. We have, after all, for the last 8,000 years been selecting among sheep for tractability, slaughtering the most aggressive ones who broke out of the corral first. How dare we then turn around and slander a species for some combination of normal herd behavior and for precisely those characteristics for which we have selected for 8,000 years? It's, I am indignant on behalf of sheep. In turn, we have been as shaped by our domesticated animals as by our domesticated plants. We prepare corrals, mangers, bedding, and stalls for them, and when necessary, bring them fodder. We tend and protect their vulnerable offspring. We milk, shear, and medicate them. Having taken them in and transformed them, we become, in a sense, their servants as well. They depend on us as we on them, like grain farming, stock rearing, sets in motion, a whole set of routines that choreographs our day and our year. Here, we're showing the new air. That's in a magnificent tableau of the new air and their cattle, and this, I'm afraid, is a dinka, dinka and dinka cattle, and since the new air and dinka don't exactly get along these days, maybe it's inappropriate of me, but it's an incredible image of dinka cattle who are like new air cattle. Here, I want to quote Evans Prichard, and if you will think back to what Michael Pollan said about his relation to the tomato plant, you will see the precise parallel. Evans Prichard wrote in the new air, quote, it has been remarked that the new air might be called parasites of the cow, but it might be said with equal force that the cow is a parasite of the new air, whose lives are spent in ensuring its welfare. They build buyers, kindle fires, and clear crowds for its comfort, move from villages to camps, from camp to camp, and from camps back to villages for its health. They defy wild beasts for its protection and fashion ornaments for its adornment. It lives its gentle, indolent, sluggish life thanks to the new air's devotion. At this point, a full suite of civilizing domestications, fire, plants, and animals is in place. So too for this reason are the critical conditions for the concentration of resources that make sedentaryization feasible on a large scale. If you will forgive anachronistic impiety, instead of Muhammad going to the mountain, this mountain actually came to Muhammad. Fire cleared the land and reorganized its plant and animal life. It made more of those plants and animals palatable. Domesticated grains, which are only palatable when cooked, brought plant life previously gathered where it occurred into a tighter circle around the domus. And domesticated animals brought game and its products virtually to the front door. At about this time, unmistakable signs of sedentaryism appear in the archeological record, pots and granaries. I'm going to end just with a gesture in the direction of some of the themes that I hope to develop. The concentration of people and domesticated animals in grain in a single spot, of course, was a perfect epidemiological storm. That is to say, almost all of our infectious diseases are zoonoses that move between domesticated animals and human beings. Many of these diseases, including measles and mumps and diphtheria, simply did not, and chicken pox and so on, did not exist before the Neolithic concentration of large numbers of people and domesticated animals in one place. And the Neolithic resettlement cap, multi-species settlement cap also brought in its train all of those hangers-on or uninvited guests like sparrows and rats and mice who found the domus and the grain and the granary and the domesticated animals all the way down to the ticks and mites and fleas and so on. This was a completely unprecedented concentration of vectors of potential disease and it explains why the Neolithic resettlement cap was a very unhealthy place to be. Mortality rates were extremely high. These diseases over time became endemic rather than epidemic, of course, but in the short run, mortality was higher and most of these populations only grew by bringing more people in from the countryside, from outside, rather than growing demographically from its own sources. What is, however, important is that these grain civilizations did have enormously high rates of fertility and these came about by virtue of the fact that the distance between births was shortened, weaning could take place earlier and whereas hunters and gatherers normally waited four years between births, agriculturalists had waited two to two and a half years between births and the result of this was a kind of, over time, a demographic advantage of agriculturalists over hunters and gatherers such that agriculturalists became the expansive imperialist population movement pushing hunters and gatherers to the periphery. So in the long run, the expansion of agricultural peoples became an absolutely crucial function and it required both high rates of fertility and it also required the concentration of population, many of whom were captured populations since most of these early wars are about capturing population and bringing them in and settling them down so that their production and corvée labor can be used. There is, then, a kind of agroecology of the state. The early state did not invent domesticated plants, it did not domesticate animals, of course, or fixed-field agriculture or irrigation, for that matter, irrigation existed before any of these states, but these states exaggerated and amplified all of these agricultural characteristics that were necessary for the thriving and expansion of the state. There is, a state needs a kind of simplified agroeconomy in order to persist and it has a kind of elective affinity. The state is incubated in agroecological settings characterized above all by grain agriculture. This means rich alluvium or Lewis soils and this can even be in places like the Andes at higher altitudes if there happens to be alluvial fans that create a lot of arable land. It's surely striking that virtually all classical states are based on grain, including millets. A rare partial exception is the Inca state relying on maize and potatoes. History records no cassava states, no sego states, no yam states, no taro states, no plantain states, no breadfruit straits, or sweet potato states. There are banana republics, but that doesn't count. Why are grassy grain crops? Typically barley, rye, wheat, rice, maize, and millets. So closely associated with the earliest states. My guess is that only grains are suited to the concentrated production tax assessment and cadastral surveys, storage and rationing. Contrast in this respect, my favorite contrast with grain is maniac or yucca or cassava. Wheat grows above ground. It usually ripens at about the same time. It's easily gathered and stored. It has relatively high value per unit weight and volume. If the state wants your wheat, it just has to come when it's ripe and can take it. Or if it doesn't like you, it can burn you out when the crop is ripe and therefore dry. Or better yet, it waits until you have threshed the crop and it takes the threshed wheat from your granary. On suitable soil, it provides the agroecology for dense concentrations of human subjects. Cassava, on the other hand, grows below ground, requires little care, ripens in a year, and most important can safely be left in the ground. And it remains edible for another two years. If the state wants your cassava, it will have to come and dig out the tubers one by one. And then it has a cartload of tubers of great weight and little value. If we were evaluating crops from the perspective of a pre-modern tax man, the major grains above all irrigated rice would be among the most preferred and roots and tubers among the least preferred. That was pointed out to me that lots of legumes like chickpeas and lentils and peas, they can also be dried as well. And it seems to me the disadvantage of these legumes as a tax crop is that they can be picked all along. They don't have a determinant harvest like wheat or the major grains. They could just be picked throughout much of their life cycle. And therefore it's very hard for the kind of tax man to ascertain and assess a harvest of any of these legumes. It follows, I think, that state formation becomes possible only when there are few alternatives to a diet dominated by domesticated grains. So long as subsistence has spread across several food webs, as it is for hunters and gatherers, sweden cultivators and marine faragers, the state is unlikely to arise. And as much as there is no readily accessible and accessible staple to serve as the basis of appropriation. Nor is it just a matter of the difficulty of dominating a population which is itinerant. The state also requires or prefers a narrow band of subsistence food, fixed fields and cadastral surveys, concentrated population. And again, rice, irrigated rice is absolutely the best for concentrating population. Historically, I think it often requires at least a large portion of an unfree population. And if it can extend irrigation, it then can control access to water and therefore increase its control over the population. If we compare this to the wetlands agriculture or the wetlands subsistence of the southern Mesopotamian alluvium, there's such a wide variety of wild plants, fruits and nuts and berries, fish and game, migrations, shellfish, small mammals that it's almost impossible for its production to be assessed and to also be taxed and appropriated. And the same is true for slash and burn cultivation as well. So it explains, I think, why the wetlands subsistence of the southern Mesopotamian alluvium was a terrain for sedentarization and population growth but not of state formation until much later. And that swidden cultivators, let alone hunters and foragers, are not a population in which one can expect state formation to take place. So the state has a kind of agroecology which it tries to amplify because it's the agroecology in which it thrives to collect people, to open new lands, to extend irrigation. The last theme that I hope to develop as well is that I think there's a period of the early states when the early states are actually quite weak and the people outside the state are still very numerous. And it's a period that lasts for a greater or lesser period of time, depending on which part of the world we're looking at. And I call this the sort of golden age of barbarians. It's in a sense that period in which very weak states are prone to, and sedentary settlements, are prone to raiding, kidnapping to the seizure of their grain and valuables in which they are more likely to be paying, if you like, a tribute to more mobile pastoralists or hunters and gatherers who can prey on them because the growth of sedentary communities is rather like one-stop foraging, if you like, for mobile barbarians. And until the states become quite strong, it is a period in which states are often not safe, they disintegrate very frequently and it's difficult to have them grow again. I think I'll shall stop there. Thank you very much.