This Week in Virology with Vincent Racaniello, Ph.D. Episode 39: Virus Structure With Vincent Racaniello and Dickson Despommier Aired 5 July 2009 http://www.twiv.tv/2009/07/05/twiv-­‐39-­‐virus-­‐structure/ Vincent: Hello everybody, it’s Vincent Racaniello. It’s a Sunday and as I was traveling last week I wasn’t able to record TWiV at the usual time. Dick Despommier is still traveling and this weekend Alan Dove is moving so he wasn’t available. So it’s just me. And what I decided to do is give you a little bit of myself and Dick talking about basic virology. This is an idea we thought of some time ago. We thought we would have small segments, 10 to 15 minute segments, in TWiV episodes where we discuss fundamental concepts about viruses. Maybe you could call it Virology 101. [0:01:00] So we recorded one on virus structure just a few weeks ago and I am going to play that for you now. So it is me and Dick talking about virus structure. And when that’s done we will come back and I’ll do some reader e-­‐mails and give you some picks of the week. So I will see you on the other side of virus structure. [Virus Structure] Vincent: Now we’ve always wanted to have some basic episodes or sessions on virology. Dick: Right. It’s to bring me up to speed I presume. Vincent: In the old way with Dick and I as when we started TWiV it was just you and I. Dick: Yep. Vincent: Let’s talk about some fundamental aspects and I’ll be the professor and you can ask questions. The first aspect that we should touch on which is very important and really influences all other aspects of virology is structure. What do viruses look like? 1 Dick: Yeah. Vincent: How do we know what they look like? I am not supposed to ask the questions. Dick: Oh yeah we can ask because I know some of these answers but…. Vincent: As you know Dick, I give several courses on viruses. [0:02:00] Dick: I do know this and I should have taken all of them. I feel guilty about that. [Form Follows Function] Vincent: And in my course this is the structure lecture is actually the third lecture. I have others, a few others to begin with, but the first slide of the structure lecture shows a house designed by Walter Gropius. Dick: Oh, interesting. Vincent: Now why do you think I would start off a virus structure lecture with Walter Gropius? Dick: Well you’d have to look at his house to find out the answer. Vincent: Because he said something which is very relevant. He said, “In order to create something that functions properly—a container, a chair, a house—its essence has to be explored for it should serve its purpose to perfection, i.e. it should be durable, inexpensive, and beautiful.” Dick: Wow. Vincent: Do you know what that school was called? The form follows function school. Dick: You Bet. Vincent: And viruses are built as Walter Gropius would build them, perfect. [0:03:00] Dick: Right. Was he part of the Bauhaus movement? Vincent: He was. He was. Dick: Well there you go, see. 2 Vincent: And viruses by definition are part of the Bauhaus movement. So virus structures are apparently very diverse, but they can be simplified. We like to simplify things. It is called the reductionist approach. Dick: Exactly, it is. [Helical] Vincent: The most simple viruses are those, structurally, are those that infect plants. In fact do you remember the first virus ever discovered? Dick: Tobacco mosaic virus. Vincent: Tobacco mosaic virus. It’s nothing but an RNA wrapped in protein. Dick: Right. Vincent: It’s a single RNA and a single protein which binds to the RNA and is repeated. It coats this protein, this RNA. Dick: Right. Vincent: This is called helical symmetry. So these genomes are called, they say they have helical symmetry, one protein coating the genome. And that’s all that tobacco mosaic virus is. Dick: That’s amazing. Vincent: It is a piece of RNA coated with protein. Dick: Are there other viruses like tobacco mosaic in that they’re helically wound around either DNA or RNA? Is there a family of tobacco-­‐mosaic-­‐virus like viruses? [0:04:00] Vincent: Yes, there are other similar viruses in plants but you know there are no similar viruses of animals. So there are viruses whose genomes are coated in protein and form a nucleocapsid just like TMV [tobacco mosaic virus] but it is not naked. TMV is a naked nucleocapsid as we call it. Dick: Right. 3 Vincent: Sendai virus, which is a paramyxovirus, rabies virus, they both have genomes wrapped in protein as is the TMV genome but they also have an envelope around them. Dick: Right. Vincent: So we think this is because in a mammal… getting from host to host needs… we need to protect the RNA. The TMV is probably brought from plant to plant by mechanical means—
farm equipment, farmers, maybe insects. Dick: Insects, lots of insect vectors. Vincent: And they are probably protected but the animal viruses are not. Dick: So the envelope comes from the animal host’s cell? [0:05:00] Vincent: Yes, it does. Dick: Ah ha. Vincent: And when we get to that step of the replicative cycle you’ll see exactly how it gets there. But, it comes from the host cell. So you can think of rabies virus as tobacco mosaic virus with a membrane envelope. Dick: What about bacteriophage, no envelopes? Vincent: Some don’t but some do. Some actually have lipid components. Dick: Really. Vincent: Yeah, not like animal viruses because they don’t have that kind of a membrane but they do have lipid components. Dick: Right. [Envelopes] Vincent: So this brings us to the second general class of virions—those that have envelopes. Dick: Ah ha. Vincent: And they are, as you said, they are all derived from the host cell. An envelope is our name for a lipid bi-­‐layer. Dick: Uh huh. 4 Vincent: And in the lipid membrane there are proteins. Dick: Right: Vincent: So the famous HA and NA of influenza are stuck in its envelope. So influenza is an enveloped virus. It has ribonucleoproteins very similar to the ones we’ve talked about for TMV except they are in pieces, they’re segmented. So for influenza there are eight such pieces in an envelope. [0:06:00] Of course an envelope, a lipid membrane by itself doesn’t have a lot of rigidity so most of these viruses with membrane envelopes have below the envelope a viral protein that gives it some kind of structure—influenza viruses have an M protein, rabies viruses have another protein under there, retroviruses have a protein—because an envelope itself is not enough. Dick: Right, but if you took away the virus component from the envelope and put an antigen inside you’d have a liposome. Vincent: Yes, that’s correct. Dick: Which is a way to deliver antigens to cells that make antibodies? Vincent: Sure, absolutely. And as we’ll see later you can make particles sometimes just by expressing in cells that protein that we’ve been talking about that give the envelope rigidity. Dick: Right, so did we invent liposomes based on our knowledge of viruses? Vincent: I don’t know the answer to that. Dick: Because if I were studying viruses that would have occurred to me. Vincent: You know when you break open cells you get vesicles also, called microsomes. [0:07:00] Dick: Sure. Yep. Vincent: So that may have been part of the inspiration. Could be that viruses, you know little packages of antigens basically, inspired it. Dick: Right. Another question that I have based on this knowledge is that can you create your own virus by creating a liposome around any viral genome—take two dissimilar parts, assemble them together and make a new virus as a result? 5 Vincent: I don’t know if you could get the genome in there. Yes you could. We know for example you can express what is called the gag protein of retroviruses, only the gag protein in cells and you will get lipid enveloped particles bud off the cell. Dick: Wow. Vincent: So if you could find a way to get something inside them. Dick: Right. Vincent: Retroviruses have signals on their genome so that they are put into the particles. So you can take those signals and put them on another sequence and get it into the particle, yes. Dick: What about infecting the same cell with three different viruses? They all have envelopes. Do they exchange… does each one have a unique envelope of do they all have generic envelopes? [0:08:00] Vincent: Well if you infected with three different influenza viruses they would exchange their…. Dick: No, I didn’t mean…. Vincent: Different viruses? Dick: Yeah, completely different viruses. Vincent: No, that doesn’t happen, there’s some incompatibility. Even if you could get a cell that would be infected by all three you would not get new viruses out. Dick: I see. Vincent: There are many problems, part of the problem is that they would interfere with each other in replication, then the packaging mechanism—that is the way to get genome into the viral capsid—is not compatible among viruses. Dick: Okay, we’ll get to that I’m sure. I just wondered how generic this whole concept was. [Capsid] Vincent: Now I am using the term called ‘capsid’ which we haven’t defined. Do you know what a capsid is? Dick: I do actually because I have had a basic course in this sometime ago but why don’t you refresh my memory. 6 Vincent: A capsid is the protein shell that surrounds the genome. Dick: Right, that is encoded for by the virus itself. Vincent: Right, now influenza virus technically doesn’t have a capsid. It has a nucleocapsid which is the RNA plus the protein, but the whole virus has an envelope. [0:09:00] Dick: Right. Vincent: And when it has an envelope you don’t use the word capsid. Dick: Uh huh. [Icosahedral] Vincent: So let’s talk about the third type of structure—just a protein shell—which is a capsid like polio virus. It has a protein shell and inside it is the RNA. Dick: Right. Vincent: Many viruses are built this way and they are built on the principles of icosahedral symmetry. All that means… do you know the geometric shape that is an icosahedron? Dick: I do. I’m a big fan of Buckminster Fuller. I presume he would have been a virologist had he not been a designer. Vincent: An icosahedron of course is a solid with twenty faces, each of which is an equilateral triangle. In fact most viruses which are made of pure protein are built with icosahedral symmetry. Dick: That’s amazing. Vincent: No other kind of symmetry. And we think it is because it’s the best way to build a shell with the fewest number of components. That’s what viruses want, they want genetic economy. Dick: Sure. Vincent: So they build these nice shells with icosahedral symmetry. [0:10:00] 7 Dick: Is that a self-­‐assembling unit? Vincent: They are self-­‐assembling in most cases. Some of the more complex viruses need some help, chaperones and so forth to help assemble, but for the smaller viruses like polio, SV40, if you express the proteins they will assemble into shells. Dick: Wow. Vincent: Polio, rotaviruses, adenoviruses, SV40, polyomaviruses these are all viruses made up of just a shell or capsid and it is built with icosahedral symmetry. Now there is another kind of structure where you take a capsid and then you put a lipid envelope around it. Now the capsid is actually technically a nucleocapsid because it just surrounds the genome and you’ve got another structure on the outside. It’s a little bit technical, but just so you know what these terms mean. And yellow fever virus is an example of that. Dick: Right, the flavivirus group. Vincent: The flaviviruses, alphaviruses, all the togaviruses are icosahedral with an envelope around the outside. [0:11:00] Dick: Interesting. Vincent: So influenza viruses have an envelope but the genome is helical nucleocapsid. Then we have a whole set of viruses with envelopes that have icosahedral nucleocapsids. Herpes viruses have an icosahedral nucleocapsid and an envelope on the outside. Dick: Vince, what’s between the lipid bi-­‐layer outer coating, this lipid layer, and the icosahedral? Is there any fluid or material in between? Vincent: In some case there is material. So for the simpler viruses, like yellow fever, the viral glycoproteins that are embedded in the membrane actually pass through the space between the membrane and the capsid and they touch the capsid. There are interactions. 8 For larger viruses, like herpes viruses, that space is actually filled with protein. It’s called the tegument and there are actually…. Dick: Sounds like a worm. Vincent: I don’t what the word means. Do you know what the word means? [0:12:00] Dick: Sure, of course I deal with it all the time. Vincent: It is used in worm biology? Dick: And insects also. The exoskeleton is referred to as the tegument. Vincent: So it has many important proteins some of which are transcriptional activators. When the virus infects a cell these proteins go in the nucleus and stimulate the transcription of viral genes. Dick: Wow. Vincent: Because they couldn’t be transcribed otherwise because the cell can’t do it. So yes, in some cases there are proteins in there. And mimivirus is an example of icosahedral virus it just happens to be very big without an envelope. Dick: Very big. Vincent: But it does have these funny hairs on the surface or protrusions that are quite long. We talked about it once before. It is basically an icosahedral capsid with these protrusions coating it. It is almost as if you took a ball and had the worms sticking out all around it. It’s surrounds it. We don’t know what they are there for. Dick: What does a mimivirus infect by the way? Vincent: Amoebae. Dick: Ah. Oh yeah that’s right we’ve talked about Proteus vulgaris. That’s right. [0:13:00] Vincent: Yeah, the amoebae in water towers, lakes and so forth. It may infect us. Some people think it causes pneumonia but we’re not sure. [Complex] 9 Then the last type of structure, see we’ve only have a handful, are what we call complex viruses because we really don’t understand them and those are the poxviruses. Dick: Oh. Vincent: They are not icosahedral. They are very big. They have a membrane, it’s a very complex membrane but it’s not a membrane around icosahedral nucleocapsid, it’s not a membrane around a nucleocapsid, a helical nucleocapsid as we described. What’s inside is very complicated and we’ll have to show a picture. So we should say that we’ll have some pictures of these viruses online to go along with this, right? Dick: Sure. Vincent: And you can go… we’ll have an example of tobacco mosaic virus, the helical nucleocapsid. We’ll have an example of enveloped nucleocapsid viruses like flu. We’ll have icosahedral viruses, enveloped icosahedral viruses and pox viruses. [0:14:00] Dick: They are very pretty when you see a lot of them together clumped up and sort of crystalized under the electron microscope with negative staining. Vincent: Very beautiful. Dick: I’ve seen them. Vince I have a question also, and that is has anybody lined up the viruses that are most widely known in terms of, on one side of a page here’s the shape of the virus, in the middle here’s the sequence for the genome, and on the right-­‐hand side here are the number of proteins encoded for and here their sequences and their structures. Has anybody ever put that all together in one page so that you can get the complete physical story of a virus just by looking? Vincent: That would be a big picture. Dick: It certainly would. Well some of these viruses don’t produce that many proteins do they? Vincent: Some of them are small. 10 Dick: What is the minimum number of proteins produced by a virus? You told me one. Vincent: One. Dick: Well that’s it. Vincent: Well here I have a picture from our textbook. Dick: Okay. Vincent: It’s not everything you asked for but it’s…. Dick: Close enough. Vincent: Have a look at that see if that starts…. [0:15:00] Dick: It starts my mind working. How’s that? [Virus Visualization] Vincent: Isn’t it true that the way you assemble information visually can be very important? Dick: In fact that would lead me into… that’s right, in fact I brought a book over for you Vince that actually helps you do that and it’s called Visual Explanations by Edward Tufte. In fact he has four of these books. I just brought you one of them because I bought an extra copy. The first book he ever wrote was called Envisioning Information. And how you envision information, I think we’ve gotten spectacularly good at this in molecular biology and you know I think of things in terms of these DNA chips and looking at the arrays and the expression of gene patterns as mountains and valleys. We’re extremely visual in terms of our ability to learn things. Our optic lobes are bigger than any other part of our brains in terms of receiving information which is why pictures speak so much louder than words. [0:16:00] In fact that’s why in my early career I was an electron microscopist. I enjoyed looking at things under the electron microscope and certainly viruses are great to look at I think. Vincent: Do you know when the first virus pictures were made by EM? Dick: My guess is back in the 50s but I don’t know because I’m thinking of the history of the electron microscope. 11 Vincent: In 1940, Helmut Ruska. Dick: Really, 40, 1940. Wow. Vincent: First pictures of virus particles, bacteriophages. Dick: 1940? Vincent: Yes, that was the beginning. Then we have lots of pictures of various viruses. Dick: Yes, they are fantastic pictures. Vincent: Fantastic. And now of course we can do structural determinations and get even more beautiful pictures. Dick: And chemistry too. Vincent: That’s fabulous. Virus structure is a huge field. I was just on a study section, a virology study section, last week and the number of applications to do structure is amazing. Dick: No kidding? Vincent: X-­‐ray crystallography, nuclear magnetic resonance, and cryo-­‐electron microscopy. Dick: Oh yeah. I’ve done that actually. [0:17:00] Vincent: That, which is where you freeze a virus and then you take pictures. You take many, many pictures. It’s basically like a CAT scan. And then you take all the pictures and assemble them by computer. That’s getting even more powerful. The resolution is getting better. Dick: You can make three dimensional models. Vincent: You can make three dimensional models. So what you do Dick is you place virus particles on a grid and you freeze them. And you take hundreds of pictures. The assumption being is that you are going to get every possible orientation. And then a computer assembles them into a three-­‐dimensional object. Dick: Cool. That’s very cool. Vincent: So in a person when you take a CAT scan the imager rotates around the person. But here the virus is rotating on the grid. Isn’t that fabulous? Dick: It’s the best. 12 Vincent: So that’s virus structure. So any other questions about virus structure before we end this session? Dick: Um, gee Vince I think you’ve covered everything I would have asked. Vincent: Alright, there will be a quiz next week. Dick: Great, I won’t be here. I am auditing this course I want you to know. Vincent: Alright that’s it for virus structure. Dick: That’s great. [0:18:00] Content on This Week in Virology (www.twiv.tv) is licensed under a Creative Commons Attribution 3.0 License. Transcribed by Steve Stokowski. 13