^B00:00:00 >> From the Library of Congress in Washington DC. ^M00:00:04 ^M00:00:18 >> Dan Turello: Good afternoon, welcome to our program, the origins of the RNA world: a collective oral history. We're glad you're here, we're glad the Metro shutdown yesterday and not today. This is a good thing. I'm Dan Turello, I'm on staff of the Kluge Center and before we begin, I want to remind you of a couple of items, we’'re recording today's conversation for future placement on the library's YouTube and iTunes channels and also please turn off any cell phones or any other devices that might interfere with the conversation. This afternoon's conversation is being moderated by Nathaniel Comfort. He is the Blumberg NASA Library of Congress chair in astrobiology. I'm going to introduce him in just a few moments, but first a few words about the Kluge Center and about the Blumberg program as a whole. The Kluge Center was created 16 years ago thanks to a generous gift by philanthropist John W. Kluge. We have two primary roles, the first is to create a scholarly community of approximately a hundred scholars every year. These are senior scholars, postdoctoral fellows, pre-doctoral fellows, all of whom contribute to a vibrant community here on Capitol Hill and in the library. The second function is to administer the Kluge prize. This is awarded by the librarian of Congress, it's a one million dollar award that recognizes lifetime achievement in the study of humanity. It was last awarded to philosophers Jürgen Habermas and Charles Taylor this past summer. So within the Kluge Center, the Blumberg chair in astrobiology is the result of what has been a unique and rewarding collaboration between NASA and the Library of Congress. The chair takes its name from Nobel Prize winner Barry Blumberg, he was the founder of the NASA Astrobiology Institute and also a founding member of the Library of Congress Scholars Counsel. Blumberg envisioned the creation of a chair in the Kluge Center that would focus on the humanistic and societal impacts of astrobiology. The idea was to connect scientific discoveries in the field of astrobiology with the best thinking in the humanities and social sciences. And astrobiology is a field of inquiry, in particular, is one that allows for the big questions. Things like investigations on the origins of life about which we are going to hear more about this afternoon. So placing scientific knowledge within the context of historical and philosophical thinking gives us the chance to consider questions of meaning and of value that are important to us as human beings. Nathaniel was the third chair to be in residence. Our first chair was David Grinspoon, who I see sitting here in the back. He researched a book on the Anthropocene. We then had historian of science Steven Dick, who wrote about the potential societal impact of the discovery of microbial or complex life beyond earth, and this year we've been thrilled to have Nathaniel with us since October and he is looking at the history of the genomic revolution in origin of life research. In addition to holding the Blumberg chair here at the Kluge Center, Nathaniel is professor in the department of the history of medicine at the Johns Hopkins University. He has published scores of articles and several books, I mentioned just a few of the more salient ones. "The Science of Human Perfection: How Genes Became the Heart of American Medicine" published by Yale in 2012, "The Tangled Field: Barbara McClintock's Search for the Patterns of Genetic Control" published by Harvard in 2001, he' also the editor of the volume "The Panda's Black Box: Opening Up the Intelligent Design Debate" published by Hopkins in 2007. He writes extensively for the Atlantic, the Nation, the New York Times book review, and other outlets. He also blogs and so you can look this up. His blog is genotopia.scienceblog.com. So we've been thrilled to have him with us this year and we'’re grateful to him for bringing such a distinguished group of panelists to the library today and we look forward to a fascinating program. ^M00:04:45 ^M00:04:53 >> Nathaniel Comfort: Well thank you Dan. It's a real pleasure to be here and I would just like to begin by, with a word of thanks to the Kluge Center. This is an absolutely marvelous and a unique place to be, just tell you how great it is when I got my acceptance letter, it had a line in it saying that ‘we expect you to protect your time for research and scholarship and reflection’. Now how rare is that, I mean in these days, so this is a wonderful place and I'm delighted to be here. And I'm particularly thrilled to have this distinguished panel of guests with me today to talk about the RNA world, which is an important aspect of research on the origin of life. And I'm just going to introduce them briefly in alphabetical order, first W. Ford Doolittle, who's from Dalhousie University in Halifax, Nova Scotia. He got a PhD in, his PhD in biological sciences from Stanford, he then did postdoctoral work with the distinguished scientists Sol Spiegelman and Norm Pace. He then joined the department of biochemistry and molecular biology at Dalhousie in 1971 and he's been there ever since. He has won a Guggenheim Fellowship, he's a member of the fellow, a fellow of the Royal Society of Canada, a member of the National Academy of Sciences, and on and on, many distinguished honors. And his research is on, has been on the molecular genetics of microbes and things like lateral gene transfer instead of hereditary transfer of genetic material across species, selfish DNA, gene structure, particularly introns about which I'm going to say a little bit in a moment, and the tree of life. And one thing that's really quite distinctive about this scientist is that he, right now, has two postdoctoral fellows who are philosophers of science, so he really bridges the sciences and humanities which is something that I find really appealing. George Fox is the Moores professor of biochemistry and biology at the University of Houston, he got his PhD in chemical engineering from Syracuse, he's the co-discoverer with Carl Woese, of the archaea, one of the three fundamental domains of life which really overturned the way people thought life evolved. He is a fellow of the International Astrobiological Society, member of the American Academy of Microbiology, fellow of the American Academy of the Association of Science, and his research has been on the evolution of the machinery of genetic translation, how information moves from nucleic acids into amino acids and forming proteins that form the enzymes and interesting structures in the body. Ray Gesteland is the emeritus professor of biology and the former vice president of research at the University of Utah, he’'s a Howard Hughes Medical Institute investigator,’ his PhD was from Harvard where he studied with James Watson. He did postdoctoral work in Geneva, Switzerland and joined Cold Spring Harbor Laboratory in 1967, staying there until 1978 when he joined the University of Utah. With Ray White, he began a new and now very distinguished Institute for Human Genetics, and since 1972, he has collaborated with John Atkins on the phenomenon of genetic recoding. He is the editor of the volume "RNA World" so he wrote the book, or at least edited it, and, which is now in its fourth edition so it's really the sort of Bible of this field, so it’'s really a delight to have him. And Walter Gilbert is the Carl Loeb emeritus professor at Harvard, he has a doctorate in physics from the University of Cambridge, and was appointed to the Harvard physics department in 1959, he then switched from physics to biology and he ran a lab jointly with James D. Watson of the double helix fame. He won a Nobel Prize in chemistry in 1980 for the invention of a method of DNA sequencing, and he's a pioneer in biotechnology, he founded Biogen, one of the first biotech companies, if not the first, he oversaw the development of a number of groundbreaking products, including alpha interferon and hepatitis B vaccine, and he also cofounded the large biotechnology company, Myriad Genetics. ^M00:10:11 Other honors besides the Nobel Prize include Louisa Gross Horwitz prize, the Gairdner award from Canada, the Lasker award, the National Academy of Sciences, he is a foreign member of the Royal Society, British Royal Society, and on and on. His research has included the identification of messenger RNA, the lac repressor, a fundamental component of the lactose gene in bacteria model system, pioneering work in recombinant DNA, the first genetic engineering, including the expression of insulin in bacteria, he is a founding, a pioneer member of the human genome project, and he coined the term RNA world. So, it is an absolute thrill to have these gentlemen here with me today and to get into a conversation with them. And before we start that, I just want to give a few remarks just to make sure we're all on the same page, just introduce you to the idea of the RNA world, alright? So I can't see, oh here we are. So what is the RNA world? Well beginning in 1953, James Watson, Francis Crick, aided by Maurice Wilkins and Rosalind Franklin, of course, solved the double helical structure of DNA. Now the double helix, the shape of the molecule usually gets all the credit, but the really interesting thing about it is those bars on the ladder crossing the spiral staircase because those are, those symbolize the nucleotide bases that pair up, so it's really two half spiral staircases, right, each specifying another nucleic acid and they realized that in the sequence of those bases was genetic information, okay? So it was in the DNA that the information was held for making proteins. DNA information goes to protein the A’s, C’s, G’s, and T’s of DNA turns into the amino acids that are strung together to form proteins. Proteins are crucial molecules in the cell, they do all sorts of work, they are enzymes most importantly, but they're also structural and the basis of neuronal transmission and structural elements in the cell, and so forth, crucial, crucial components of the cell. And in 1956, three years after the double helix, Francis Crick coined what he whimsically called the central dogma of molecular biology and Watson drew it out in this cartoon, which says a number of things about the flow of genetic information. This is really the first time people began to think about genetic information, and the important thing that most people remember of the central dogma is this simple phrase ‘DNA makes RNA, makes proteins, okay, so what's DNA versus RNA? They're very similar molecules, there are really only two differences between them, one of the four bases is different, RNA uses Uracil instead of Thymine, and you'll see that there's on the ring, there's one difference, an OH instead of an H, those are the only differences between the two molecules but those two slight differences make huge differences in the chemistry, and DNA of course forms a double-stranded molecule. RNA is usually single-stranded, and that means it can bend around and form into all kinds of different shapes, alright? It's also much more reactive than DNA, and those two facts become important in the RNA world. Okay. So the DNA makes RNA, makes protein through two processes, one called transcription where an RNA molecule messenger RNA, which Dr. Gilbert was a key figure in identifying, and then that messenger RNA is translated into a sequence of amino acids that makes a protein, okay? And as I said, proteins are ubiquitous in the cell, they form hair, nails, muscles, nerve cells, and most importantly enzymes. There are a couple of, and that translation process is really sort of the heart of what we’'re going to be talking about today. Translation occurs in structures called ribosomes, so the messenger RNA is read off the DNA, and then it moves outside the cell nucleus to the outer part of the cell and in the ribosome it is pulled through like a tape through a tape recorder and it is read off and the sequence of those nucleotides specifies the sequence of amino acids so this building chain of amino acids then forms the protein. Now you'll notice a couple of things here, one is that there is an awful lot of RNA here, you have the messenger RNA, ribosomal RNA, the ribosome itself is made of RNA mostly, and then, I've got one more, and then transfer RNA is the molecule that actually attaches to the amino acid and brings it to the building chain. So there's a lot of RNA in this part of the cell, okay? People very quickly began to notice this fact and in 1962 one of the pioneers of molecular biology Alex Rich gave a paper on the origin of life so this was just nine years after the double helix and in fact was the year that Watson, Crick, and Wilkins won the Nobel Prize, and he said the hypothetical stem or parent nucleotide molecule was initially an RNA-like polymer which was able to convey genetic information as well as organize the amino acids into a specific sequence to make proteins, okay? So there's a, so RNA does most of the work of that process. And in 1965, just 3 years later, two other scientists made similar realizations. J. B. S. Haldane said that life, by which he meant indefinite replication of patterns of large molecules, can be based on RNA without DNA, and Fritz Lipmann at the same conference said that DNA probably evolved later than RNA, so there was a time in which you had RNA and proteins and not yet DNA. So back to this diagram here, another thing that we can notice about this is that there are enzymes or proteins involved in each of these processes, so none of this, although there are RNA molecules involved, none of this can happen without enzymes, without proteins, alright. So these things are specifying proteins and you need proteins to do it. So that leads to a kind of chicken and egg problem, alright, where you have genes that store information, that encode proteins that make enzymes and you need those enzymes to catalyze the reactions to copy the genes. So how do you get out of that problem? The RNA world is the way you get out of that problem, okay? And several people noticed this in a speculative way in the late 1960s. Carl Woese said that proto-RNA could have been, the proto-ribosomal RNA, could have been the original genome, and Francis Crick the next year said that possibly the first enzyme was an RNA molecule with replicase properties, the ability to copy other RNAs. Thus a system based mainly on RNA is not impossible, and the same year his colleague Leslie Orgel said, asked could polynucleotides of RNA with well-defined secondary structures folding act as primitive enzymes? Could they be more than just information storage molecules? Could they actually catalyze reactions too? That would be a way out of the chicken and egg problem. So the answer is yes! But hang on one second. First I want to tell you about a couple things that are going to come up in the conversation. One is the, as I mentioned earlier, the discovery of the archaea, this brand-new group of single celled organisms that George and Carl Woese recognized through sequencing RNA, ribosomal RNA, were different from the bacteria, and so this created a lot of discussion about how, what was the original branching structure in the tree of life. ^M00:20:23 And the same year, 1977 was a big year for origin of life research. Phil Sharp and his colleagues at Yale and Rich Roberts at Cold Spring Harbor and his colleagues independently discovered that genes come in pieces. In higher organisms, eukaryotes, organisms with a membrane-bound nucleus that contains the chromosomes, have genes that can be segmented, okay, they can, and when you go through the transcription and translation process, those middle bits between the segments get spliced out and the segments join it together to form the final RNA that gets read into the protein. So that's really odd, genes come in these pieces. Walter Gilbert wrote an article the next year called Genes in Pieces and in that paper he named those segments exons, for the segments that are part of the genes, and introns as the segments between the exons, so they're inter, in between the exons, okay. And he and Ford Doolittle over the next few years got in a lively discussion about which came first, when introns were invented evolutionarily. Were they part of the very first organisms or did they get invented later as the eukaryotes evolved? So this was a very important discussion in origin of life research that I want to talk about more later. And then finally, the discovery of RNA enzymes. Crick and Orgel were right. RNA can act as an enzyme and that was found first by Tom Cech and, in 1982, and then Sidney Altman at Yale found that a molecule called RNase P that he'd been working on for years also had, the RNA also had catalytic properties. So RNA can act as both a catalyst and an information molecule. So that breaks the chicken and egg problem, alright? So this is kind of like the old Saturday Night Live sketch where you have the, you know, the shimmer which is a floor wax and a dessert topping all in one, right? The next, just a couple of years later, Walter Gilbert wrote an article called The RNA World, and this was the coining of the term and we're going to talk more about that I just want put that up there so you have a sense of the dates, okay. In 1987, Cold Spring Harbor held a meeting on RNA catalysis that involved a lot of discussion about the RNA world, and in 1993, Ray Gesteland and John Atkins edited a volume of, a collection of papers called the RNA world, and as I said, it went through multiple editions I forgot to put up the fourth. What year did the fourth edition come out? 2011, okay, so this is still a very vital and ongoing project, alright? So that is a quick introduction to the RNA world. Now I'm going to sit down, I'm going to mostly turn it over to the them. Thank you. Okay just wave your hands or something if you can't hear or can't see. I’m trying to get out of the way but I also want to make sure that I'’m in conversation with my guests here. So, with that, let’'s begin with evolution. Theodosius Dobzhansky famously said that nothing in biology makes sense except in the light of evolution, but the early generation of molecular biologists were often criticized for not taking evolution sufficiently seriously. One of those critics, at least in my reading, was Carl Woese. What do you think? Is that criticism fair? To what extent did the RNA world emerge from evolutionary thinking, I mean from thought based on serious reading of evolutionary literature. >> Dr. Walter Gilbert: I thought that, I think it’'s an unfortunate question. I do agree the early generation of molecular biologists did not, most part didn't think of evolution. The discovery of the DNA sequencing in 76, discovery of DNA sequencing in 76 suddenly made it possible to look at the genes of many organisms and one of the first things that happened as molecular biologists did this was they discovered that the genes in [inaudible] were like the genes in plants and stuff like that and suddenly evolution swept the field. I remember going to a conference in which in fact this sort of suddenly became a fantastic element in people's thinking. Those of us who had a slightly more genetic background were more conscious of evolution, but the general field of molecular biology was not. I think the RNA world image is deeply based in a notion of evolution. The point of the paper that I wrote in 1986 was twofold. One, it picked up from arguments that had been made, I would say picked up from the discovery of the RNA enzymes that had been made and reiterated just before that and had picked up from an older notion from the biochemists that RNA was more primary than DNA, and the biochemistry of that is that all of the DNA structures, the DNA sugars and the DNA, of the DNA bases and nucleotides, are made from RNA precursors. So if you look at the biochemistry, you would not say DNA is separate in any way or primary in any way, you’ would say, oh, the chemistry suggests that RNA is the thing that you made biochemically and from that you’ would later make DNA. People knowing that thought of a RNA protein world. RNA is a genetic material, machines in the organism, and that genetic material can be used to dictate the sequence of amino acids and amino acids were thought of as the interesting things in the world, the enzymes. And the protein chemists ignored the nucleic acids intensely, they thought everything interesting in the world was in the proteins. It makes very interesting meetings in which the protein chemists meet separately and the nucleic acid chemists meet separately and they never like to talk to each other or the protein chemists totally ignore nucleic acids. >> Nathaniel Comfort: Nobody could translate between the two languages. >> Dr. Walter Gilbert: But the discovery of the enzymatic activity of RNA made me think that maybe RNA could be entirely, play the role of all of the enzymes and so the paper I wrote in 86 makes actually two statements. One, it suggests maybe RNA enzymes could in fact do all of the necessary activities for a cell to exist, including the copying of RNA itself, and therefore you could imagine evolution is beginning with a small RNA molecule that develops by random structures, random association of bases, the ability to copy itself, and once it can copy itself, it can make more, makes more copies, but in fact since it'’s a biological process, it makes copies which would also have errors and so it can evolve. It can make random changes and those random changes can be acted upon by natural selection to eventually produce better molecules. Second part of that essay, however, points out that because one of the RNA enzymes that was then known at that time was an intron, the region inside an RNA molecule, it could splice itself out. I could make the argument that that activity clearly could be used, not only to splice introns out of the RNA, but by combining two of those structures, develop a structure that can move an RNA piece around from one molecule to the other, inserting it like a transposon, splicing out the rest, and creating a novel RNA combination, and that's a very powerful evolutionary tool, we call it recombination when it occurs at DNA level, but in fact here was a model that clearly could happen at the RNA level, and so I could immediately in that paper suggest two things. Maybe there could be RNA-based organism, completely using RNA as enzymes, and the evolutionary processes would be able to shuffle the pieces of RNA around to make novel pieces again. ^M00:30:00 The original intron, exon idea you alluded to, the critical element of that idea was an evolutionary argument, it was not just the biochemical argument that you had on the DNA long regions that we call introns that are spliced out of the RNA molecule to produce small pieces, together they code for pieces of protein. But the evolutionary argument is that by spreading those regions apart from the DNA, you would increase the recombination rates between those regions and hence you could more rapidly create protein structures and evolve them to better function by recombination and by shuffling the pieces of the protein. So in fact both the intron, exon idea is conditioned by a vision of evolution and the RNA world idea is also conditioned by that vision of evolution. That paper in fact finally includes that sort of a hidden definition of life as that thing which can by replication, mutation, and recombination as by changing random changes can evolve and thus can be operated upon by natural selection to reduce ever better functioning. >> Nathaniel Comfort: Okay, thank you. I think that was a very fortunate question in that case. I can now throw away half of my questions but [laughter] Ford I think you might have something to say on this. >> Dr. W. Ford Doolittle: Yeah well, I think your initial question involved the phrase ‘serious reading of the evolutionary literature’ or something like that and I think most molecular biologists did not seriously read the evolutionary literature, probably still don’t. Mostly evolution is something you talk about in the last paragraph of the paper and I think, and over a few beers kind of thing because I think there was a general feeling amongst the molecular biological community that evolution obviously was important but also there is nothing you can really say about it so you could just say whatever you wanted and everybody would accept that politely. >> Nathaniel Comfort: I think that’s right. >> Dr. W. Ford Doolittle: I think you were right that 1977 was a very important year for some of us and I want to go back a little bit about the intron relationship to the RNA world because I wasn't always lab on sabbatical at that time and [inaudible] came back from Switzerland, I think, having heard for the first time about the introns in the immunoglobulin genes, I believe, and gave, there were live meetings once a week in all these labs with very strong tea as I recall. >> Nathaniel Comfort: Why tea and not something stronger? >> Dr. W. Ford Doolittle: And while we presented the exon shuffling idea, which I thought was a brilliant way for things to evolve more rapidly in the way that he just described, but I had one concern about that which was I believe thinking the way that most of people, for most people thought in those days, we imagined that higher cells, eukaryotes, animals and plants evolved from bacteria and bacteria don't have introns and eukaryotes, higher cells do have introns and so that sort of implied that these relative event cells would suddenly take on these interruptions in all their genes just so that they can in the future do a better job of evolving and which seem, not something could, anybody who have done any serious reading on evolutionary literature would be willing to accept is possible. >> Nathaniel Comfort: You're not quite fair. >> Dr. W. Ford Doolittle: I'm not blaming you for that. >> Dr. Walter Gilbert: I was going at large, at that moment we knew of introns only in more complicated genes, and so the first suggestion, I learned about the introns and exons from the people who discovered the splicing in the Cold Spring Harbor meeting in probably May or June that year and in my laboratory we had a sequence of an immunoglobulin gene which actually had an intron and we just sequenced it. We didn't actually know what that was, but it was an intron. So when I came back from Switzerland having thought about these ideas, I suggested that eukaryotes uniquely use the greater evolutionary speed of recombination within introns to reduce the great explosion, pre-cambrian explosion of the eukaryotic genes, and that'’s the background. >> Dr. W. Ford Doolittle: And that’s what I objected to. >> Nathaniel Comfort: Right. >> Dr. W. Ford Doolittle: Because of, if we believe, which I think 99 percent of the people believed at that time that eukaryotes came from prokaryotes, then they would have had to take on the burden of introns and ten introns on average on every gene in order that several million years down the road they’'d be able to do these wonderful evolutionary things which seem kind of anticipatory and not the kind of thing, evolution really can't look ahead so that was my reaction to what you, and I don't think you actually implied that but that'’s my reaction. And I also happen to be privileged because I had worked on [inaudible] and I knew Carl Woese and I knew George and I knew about this almost simultaneous publication of the three the main view of life that George and Carl Woese had been working on that, in their view, eukaryotes and prokaryotes were not evolved, I mean it wasn't that eukaryotes evolved from within the prokaryotes, they evolved separately from some more primitive ancestral form, so then I thought, well actually went home and had a bottle of scotch and wrote overnight that piece and I thought that, you know, it'd make more sense if introns were actually present from the very beginning. And then as evolution proceeded, prokaryotes lost all the introns but eukaryotes retained them and this gave them the evolutionary ability to become complex organisms like ourselves and prokaryotes have'n’t done that although they've done a lot of other wonderful things and they've gotten much more streamlined, we use the word. >> Nathaniel Comfort: Streamlining, yes. >> Dr. W. Ford Doolittle: