9UNP0435SD_Bret_Payseur

- And welcome to tonight's Wednesday Nite at the Lab. We're having Dr. Bret Payseur. He received his bachelor's degree in molecular, cellular, and developmental biology and anthropology from the University of Colorado Boulder in 1996. He earned a master's degree in evolutionary biology from Northwestern University Medical School. Bret received his PhD in ecology and evolutionary biology from the University of Arizona in 2003 and performed postdoctoral research at Cornell University. Bret has been an assistant professor in genetics at the University of Wisconsin Madison since August of 2005. Bret uses genomics to understand the evolutionary processes that shape genetic variation within and between species. Everyone, let's welcome Dr. Payseur tonight. 

(applause)

- Bret Payseur: Thank you very much. Is this sufficient volume for everyone? Can you guys hear? Sort of? More? What about that? Is that better? I'm asking kind of quietly. Thanks a lot for the invitation to speak here. 

This is really a privilege to talk with this group. And I wanted to mention a few things just for fun. One is, some of you might know this because of your connection to the university, we just celebrated, in the Genetics Department, our centennial this last weekend which is very exciting. So another reason to take pride in the University of Wisconsin is that we have the oldest genetics department in the country. And it's still kicking. So about everything I'm going to tell you about tonight was really made possible by the Genetics Department, and I have a lot of gratitude towards them. So I have a pretty informal presentation here, and I actually haven't been to one of these before. I'm sorry. I'm sure after tonight I'll come regularly. 

(laughter)

 But feel free to interrupt me. I want you to get something out of this. I'm not trying to put on a show or anything. I'm sure you can teach me quite a few things as well. If we get into discussions, that's fine with me. However you want to do it. But let me just get started here. So I thought, we have a number of topics that we focus on in my lab, and I thought I'll just mention a few of them so that you don't think I'm limited to the topic of speciation. One of the things that we study is what is called human genomic variation. And what we mean by this is just taking the DNA of lots of humans, comparing it, and using the differences and similarities to understand something about human relationships to each other and human history, evolutionary history. 

Another topic that's not exactly related to that is the evolution of giant island mice. So there's a group of house mice that live on an island in the south Atlantic called Goth Island that have evolved to be about twice the size of the regular house mouse. And we're currently trying to figure out the genetics involved in this change, but another sort of interesting behavior is unlike most house mice in the world that prefer to eat things like seeds, they eat birds. They actually eat albatross chicks. So for those of you who know the size of an albatross, that's pretty amazing. But a topic that we spend most of the time discussing in my lab is really this evolution of new species which is what I want to talk to you about tonight. And I'm going to do this kind of in two phases. The first part of the talk I'll just sort of walk through some of the basic things that we know about how speciation happens. And then I'll show you some data from my lab that are related to that. So, why does anybody want to study speciation? 

What's the point of this? It seems really controversial and difficult to understand and we have long timeframes to look at, it's very challenging. I think the reason is that we're fascinated by biodiversity. We look around the world and what we see is enormous diversity. There's a real splendor of life forms. And there's a kind of natural curiosity about why so much diversity. You can easily imagine a planet that had organisms that only had a few types. But that's not the place where we live. And so we don't really know what this number is, how many species there are in the world. But there have been many estimates that have kind of put it at 10 to 50 million. 

Many of these have not been described, and of course there are probably many more that we just don't know about. But there's a lot of species. So if you think about biodiversity in terms of the number of species, just for a moment, if you think about it for a second, in a very basic way the number of species that we see on the planet has to reflect a balance between two opposing forces. On the one hand, we've got extinction, the loss of species, and as this extinction rate goes up, we're going to have fewer and fewer species to look at. On the other hand, we have speciation, the birth of new species, and as that increases we're going to go towards more. And there's sort of, you can think of it as this constant battle between these two forces in determining the number of species that you see in any given time. So we know quite a bit about extinction and speciation rates, mostly from looking at the fossil record. This probably won't be news to many of you, but there have been, people believe, I guess it's six sort of major extinctions that we know about during the history of life. And with a really big one that happened in the Cretaceous, and some people believe that we're sort of on the road towards a seventh, based almost entirely on human activity, but that's for another night, I guess. So the point that's interesting here is that as you look across time periods where people have measured these types of things, you see two things. 

First the extinction rate is sometimes low but it's also nonzero. There's always species going extinct at any time frame you look at. And all they have to do is open up the New York Times, unfortunately, to see descriptions of species that are going extinct as we watch them disappear. So there's an enormous sort of conservation implication here. But the other thing to notice is that there's variation in the extinction rate. Some periods of time you see more species disappear than others. And there's a lot of discussion about why. We can do the same thing with rates of speciation. And that's what is being shown here. The percent origination is just a measure of the rate of speciation, sort of birth of species in the fossil record for a particular type of animal. 

And you don't need to pay attention to all of this except to see we're just moving forward towards the present in geological time. And you see, again, there's plenty of speciation happening, plenty of new lineages being born all the time, but there's also variation in the rate at which that happens. And this is just based on looking at organisms for which the fossil record is good enough to do this. So in terms of the importance of speciation, we have the fact that it's a major contributor to biodiversity. And sort of more generally, we can think of it as a primary engine of evolutionary change. Evolution is happening in a number of different ways, but the birth of a new species is a pretty fundamental one. And, in fact, it's one of the things that sort of puzzled Charles Darwin. Something that might be a little less obvious that we could come back to later is that like most of evolutionary biology, an understanding of speciation should be quite useful in areas as diverse as conservation, agricultural, and even medicine. So we'll come back to that down the road. So hopefully you agree that speciation is important. 

Now here's the thing about studying speciation, is you have to be prepared to say what species are. And so I thought this would be sort of one way of going at it. When you look at organisms, they show up in different flavors and there are lots of different ways that you can compare them to each other and organize them into groups. 

I think we can kind of think about the definition of a species the way we would think about how to organize these chairs. And each of you can probably come up with your own concept of which ones of these should go together. I actually stared at this for a while and was amazed at how many I could come up with a way to unite every possible group and it's sort of interesting. But obviously if I was looking at the sort of style of incline, I might put these two together, but if I was looking at the materials they were made out of, this one might be put in its own group. So on and so forth. The point I want to make here is that this question about what is a species, although it determines how you study speciation, there are a lot of different ways to define a species. And I think that what biologists do in practice, even though there's been a lot of debate about what a species is, is to try to just define species in such a way that the process of how new species arise is something that can be studied. 

So it's almost like a methodological kind of approach. So there are about as many species concepts as there are people in this room that have been proposed over the years. Everybody has their favorite idea. And I will just sort of kill the suspense for you right now and say there is no species concept that encompasses all life. I don't know of one. It's very difficult to do that. So what I'm going to talk about today doesn't really apply in a very straightforward way to these somewhat abundant life known as bacteria. But it does apply to some other organisms in a helpful way. So here I'm just showing two species concepts that are probably the most popular ones among biologists. The first one is if we had a way of just reconstructing the evolutionary history, measuring the relatedness among different groups, and found that a particular group of individuals shared a very recent common ancestor. 

And how recent it needs to be is a little bit up in the air. But they share a recent common ancestor here to the exclusion of these other lineages. Some people are in of favor calling this group a species. And it's just because they have all of this sort of shared evolutionary history right here. That's the idea. It can be difficult to decide how close they have to be, but most species concepts run into that general issue. So then the other one which is probably something you may have heard of is several people argue that the lack of actual or potential interbreeding among individuals is a great way to define species. At least in organisms that are sexual. And this is coined "reproductive isolation." And I just have here a mule which most of the time is sterile. 

So under this kind of scenario, if you're going to define reproductive isolation this way, then you would probably call horses and donkeys different species. Which we do. So this is the concept that I'm going to use throughout the rest of the talk. So if you get angry about that, let me know and we'll resolve it. So once you pick a species concept like that, and these species are called biological species, the ones based on actual or potential interbreeding, then the problem becomes a little simpler. That is, we can study the mechanisms underlying this reproductive isolation, and that will hopefully give us insight into how new species appear. Because now we have something to look at. So I just want to walk you through the different kinds of reproductive isolation that show up with these biological species. And you can probably think of even more than the ones I mentioned, but these are sort of the most obvious ones. So first of all, of course, you can have populations that, over time, find themselves in geographically isolated areas. 

And this can be because one population or individual has moved over here and started a new population and over time maybe there was a flood that limited the migration between the populations. You can probably imagine a lot of different ways that this can happen. But you see populations of organisms all over the world that aren't free to mate with their neighbors several, whatever, geographic units away. So that's geographical isolation. Another thing that can happen, before fertilization again, is ecological barriers can arise. And what's meant by this is even if you're living in the same geographic location, you might have divergence and habitats between groups. So what do I mean by that? If you, for example, were looking at insects, phytophagous insects that live on plants, and you might find that there are two sort of different types of plants in a particular locality and that they're largely inhabited by different populations of insects. And over time, if that association remains, those insects will start to behave like different populations. So that's the idea. 

Another form of isolation you can get is just temporal. And here what's meant by temporal is just activity patterns. So if you start to have divergence in when a population is active relative to another one, then that's going to create a barrier to reproduction. There's a couple more. We're still talking about before fertilization. So this first one is something that's sort of obvious. We know that if we talk about mammals, which is what I'm going to talk about tonight in particular, that females are choosey. I'm pretty sure we all know that. 

(laughter)

- Choosey.

- Choosey, yes. And in mammals, females, again here's a fact you probably already know, in mammals females tend to be choosier than males do. Just saying. 

(laughter)

 So anyway the point is that individuals can have mate preferences and they certainly do. And over time, those mate preferences can be solidified and eventually lead to a situation where there isn't the possibility of mating anymore. That's the idea. There are also obvious sort of mechanical constraints that can happen if you just, I was trying to think of a picture to put here and I just decided to let you use your imagination. 

(laughter)

 But there are other things that can happen that sometimes referred to as gametic incompatibilities, meaning that the sperm and the egg don't get along. 

So there's something that's blocking fertilization even when gametes are being exchanged. So these are all barriers that you can imagine, but we also know that there's hybridization. When you look the in nature, you see things that look different that mate. And I just put a few examples up here, which I just realized are all mammals, I have a little bit of a taxonomic bias. Actually, the most frequent sort of life forms that hybridize are really plants. Plants are very good, and you probably all know that, but they're very good at tolerating hybridization. Animals are not as good for reasons that are not fully understood. So these are a bunch of examples of hybrids. I'm going to get the names wrong. Maybe some of you guys will know.

- A liger.

- What's that?

- It's a liger.

- That's a liger. So you know what that's a cross between?

- A lion and a tiger.

- Very good. This one we already talked about, it's a mule. What about this one? These names are so ridiculous.

- It's a camel and a llama.

- Yeah, so what do you think you call that? I think it's called lamel and there's cama. They're not very imaginative with some of these names.

- What's it a cross between?

- Between a camel and a llama.

- Between a camel and a llama?

- A camel and a llama. This one is a cross between a dolphin and a false killer whale. So I don't know what happens in this individual's germ line but we'll talk about that in a second. 

You guys know what this is? I don't know why you would. It's either a pizzly or a grolar. 

(laughter)

 Exactly. And they look different depending on which direction you go, like who's the mom and who's the dad.

- Can the offspring reproduce?

- You know, that's a good question. I don't think we know very much about this one, about the bears. I don't know about this one. 

It's a grizzly and a polar bear.

- Are grizzlies and polar bears different species?

- Well, this is what we're going to sort of talk about. 

(laughter)

 They might be. They might be. So the point I wanted to make here, which you nicely led me to, is that plenty of things can hybridize, and you could decide to say anytime you make viable hybrids of any sort, you're one species. But what biologists do in practice is to recognize that even if you can produce this thing, it might be at some selective disadvantage. So it might have lower fertility, it might not live as long, who knows. It's sort of up to us to figure that out. 

So I would be comfortable calling two groups species if their hybrid offspring are in some way less fit than the pure offspring would have been. That's sort of a general way of looking at it. But in practice it's kind of hard to decide exactly where to put that.

- Does it ever happen in their benefit?

- Yes, sometimes it does.

- Do you make a distinction between whether hybridization could be engineered or whether it would actually happen out in nature?

- Wow, this is a great audience. As I'm going to say in a minute, I think measuring interbreeding in nature or in the lab, both of those approaches are helpful. As I said, even if you can produce hybrids, they can be less viable than the parents and often they're sterile or at least show reduced fertility. So this is a form of reproductive isolation after fertilization, which is the one we study in my lab. 

But let me just, now that we know what reproductive isolation is, let's just talk briefly about how it shows up. How does it evolve? Because you start with a population somewhere where everybody's breeding fine. So how do you get to a point where they're not? Here I'm going to show a couple simple schematics of this so you can see what you think. So imagine you have just a population of mice, and as we said before, there's this kind of geographical isolation that happens over time. And the important point of this, these mice, some of them traveled to a different place. It's actually pretty easy to have this happen with mice because they can just hop on boats or pick-ups or planes. So they end up in different places and they can't really get back. The important point is that over time, these two lineages, if they're evolving independently, it means they're not exchanging genes anymore, then they're going to accumulate mutations, as all populations do, and over time some of them might raise in frequency until everybody or most of the animals in the population have that change. 

So here I'm just showing a coat color mutation which we actually know a lot about as a field. And so this is a simple way to evolve reproductive isolation because we just had regular old mutation and evolution, and when these mice now come back together, if they should in the lab or in nature, perhaps the female gray mouse prefers other mice that are gray. And suddenly we have some reproductive isolation. Does everybody see that? It's pretty straightforward. You just have to believe that populations can get isolated and that genetic changes occur that affect traits. So hopefully that's pretty easy to swallow. And let me just show you, sometimes people ask me what's the evidence that this actually happens? It's a little bit difficult to observe in nature. So here's a really classic example of how this happens. 

And this is an experiment done by this woman Diane Dodd many years ago. But this kind of experiment has been done many times, particularly in Drosophila. So these are fruit flies. And what she did was to just take a single population of fruit flies and then sort of separate them into two groups. One that was eating more starch-based food and another one that was eating maltose-based food. So just a sugar. And the idea was that maybe if these flies, over generations, adapt to the different food sources that some aspects of their reproduction might sort of come along with that selection. You're sort of accelerating the evolution of these flies, and so maybe it will cause them to develop some kind of reproductive barrier. So lo and behold, I think it was actually like 12 generations when she started seeing the first signs of this, but you see many changes in the flies, mostly related to the way they digest the food source, as you would imagine. So this is natural selection and a response to natural selection and action. 

And then she put them back together and lo and behold the flies that were fed on a particular food source preferred to mate with each other instead of the ones on the other food source. That's kind of impressive because it's a pretty short amount of time. So I would argue that this type of thing is happening sort of all the time in nature. You might not have selection that is this strong all the time, but the process is something that seems likely to occur. So that's kind of an example of that before fertilization type of isolation evolving. But I just want to mention real quickly that the after fertilization part, after reproduction part, we think it might operate in a slightly different way. And so if you think about this for a second, the example I just showed, which led to these two types of flies that couldn't mate with each other or didn't want to mate with each other, you really only need one change for that to happen. That's the minimum number, you need one of the types of flies to develop a preference. But if you think about the way that hybrid sterility or hybrid inviability happens, it's harder to imagine that it would happen with one change. And the reason is if I have a population of individuals and I have a mutation that comes into the population that makes them sterile, how is that mutation going to spread? 

It can't. So that's the problem. So that puzzled people for many years and then in the 1930s and '40s, a couple of people independently came up with this idea that you need two genes, you need two mutations. And this is a very simple idea. So we have this same thing happening again with our geographical isolation. So I have independent evolution. And what I'm showing here is just imagine there is some gene, the red gene, that does something, maybe related to fertility, and then it interacts in mice with a blue gene. That is, you need both of them to do something. There are lots of traits like this, genes are organized in networks, usually, so it sort of makes sense. So now imagine that we have a change in the blue gene here and it's just fine. 

It's a change but it doesn't cause sterility or inviability. So it just happens by normal evolution and comes to represent most of the population, and here we have a change in the red gene. And the important point is that even if natural selection saw this red gene, this red mutation and the blue mutation and caused them to spread through the populations, when they come back together, the combination of the red mutation and the blue mutation could cause a problem. The combination could lead to animals being sterile even though we never went through a phase where anybody was sterile. So the important point here is that it's the interaction that matters. And this is the model that most biologists believe is the cause of things like hybrid sterility and hybrid inviability. So if you look at the mule, even though we don't know what the exact genes are, most people believe that there's been a change in donkey and a change in horse that interact unfavorably in the hybrid. That's the general idea that we think of. So what do you need to evolve reproductive isolation? You need genetic changes. 

You need something heritable so that it will persist through the generations. These changes then need to be spread throughout the populations that they're in. And that can happen, as I've mentioned, either by natural selection, increased fitness, or just by chance. Mutations change in their frequency just by chance. That happens especially, that effect is especially dramatic in small populations. And then one thing I just want to mention is I'm talking about sort of the simplest models where you have this geographical isolation, but it's also true that new species can arise from within one population. And we talked about some of this before in terms of adapting to different habitats within sort of one location. This mode of speciation, excuse me, has been demonstrated to occur, but it's rare in comparison to the sort of population separation type of speciation. This is a very active area of inquiry in evolutionary biology to understand how often this happens. Yeah?

- Have you every heard of a case where there's essentially a culture origin? For example a red-feathered bird different want to mate with blue-feathered birds?

- And then that got sort of spread by word of beak? 

(laughter)

 So that can happen. The main thing is then that you sort of need some way, so yeah, that's a good point. To the extent that the culture can be transmitted in a stable way, that can happen. But it needs to sort of act like genes that way. You know what I mean? It needs to of inherited. 

Some people think that there's a lot of debate about how often that happens. But in principle, it makes sense.

- Could you give us an example of the new species from one population?

- So there are these fish, these cichlids in Africa, and, boy, this mode is tougher to explain. But, basically, the evidence that this is called sympatric speciation, by the way, which just means that it's happening and it's sort of overlapping ranges. The evidence that sympatric speciation has occurred is that when I look at the genetic diversity of these cichlids, you can see that, and you look at them in a bunch of different lakes, you can see that fish have their closest relatives, evolutionarily, first of all there's a whole bunch of species in each lake but that the closest relatives are always in the same lake. Okay? And so it's like they get into a new lake and then something happens. Now the argument that people have made over time is that well maybe within the lake they're in different environments. And that's a hard one to refute. 

But in the case of the cichlids, they swim right past each other. So they might be seeing something that we're not, they probably are, but that's kind of the most often quoted example. Yeah?

- So like Darwin's finches, would that be an example of that?

- No, Darwin's finches, the degree of sort of isolation between them is still being studied, but they certainly have some of the hallmarks of sympatric speciation, that's right. Yeah?

- I didn't quite get your explanation of the necessity of two mutations, two genes. Where there's one, you said now that would supposedly made you infertile, made the organism infertile, then how could that spread? So you said maybe it takes two genes. So now this organism has two genes that makes it infertile, but how does that spread?

- Well, the idea is that you have independent, so the sterility is caused by the combination of two mutations.

- So how does that spread?

- In these individual populations, there's only one mutation. So this population has one mutation, this population has a different one and a different gene. So those don't cause sterility. So they can just spread through the population. And then when you put them back together, they can just spread within their individual populations. That's right. And then when we come back together there's trouble.

- But how come there's trouble? 

Because the two genes together can't spread?

- Right. The two genes together don't spread, it's just individual.

- So why is that trouble? If it doesn't spread?

- Explain why there's now two species when there used to be one.

- Maybe we can revisit this in a little bit. Is that okay? I'm glad there's so much interest. This is good. 

I just want to point out, in the interest of either defusing or creating controversy, this kind of process that I just described, you can think about it in fruit flies, you can think about it in yeast, whatever you want, but the same kind of thing is what happened in human evolution. And so what I mean is you can imagine some ancestor here that lived five to six million years ago, and these are chimpanzees and humans, and at some point those two lineages split and they started to accumulate genetic changes. Some of the genetic changes are responsible for the conspicuous lack of hair in humans. Some of them are responsible for the brutish behavior of one of the species. 

(laughter)

 There are lots of changes that occurred that affect these traits that we can notice. But I just want to point out that there's no reason why, I don't know how much we know about whether human-chimp hybrids are sterile. Again, that's probably not something I should speculate about. But you can imagine this same type of thing happening where, say, there's a mutation here and one here that do fine in their individual lineages but then if you were to put them together, there's a problem. In this sense, there's nothing different about how humans evolved in terms of speciation. 

And there's lately been a lot of arguments about whether, when this split happened, whether there continued to be early on hybridization between the lineage leading to chimps and the lineage leading to humans. Yeah?

- Wouldn't there have had to be because if there hadn't been a misjunction of two primates, whatever, who were close enough and managed to breed but their offspring was sterile, that would work to maintain their separate arms, wouldn't it? Because otherwise you'd get another line coming down. One from this side and one from this side made it.

- Yeah, right, unless it didn't make it.

- Exactly.

- Yeah, that's right. So here's the thesis that I'm going to say here is that we can defuse a lot of the mysteriousness of speciation if we just think about it this way. It's a little bit of a reductionist way of thinking about it. 

But if we can just look for these genetic changes that are responsible for reproductive isolation, that tells us, to some degree, the mechanism of how we get new species. That's the idea. And that's what we do and so I'll just, fairly quickly, run through a few examples. We use house mice for this for a number of reasons. One of them is that house mice, as I'll say in a minute, are separated into three closely related species. They are closely related enough that people often refer to them as subspecies. But, as I always say, since I would prefer to be studying speciation and not sub-speciation, I call them species. But it's sort of arbitrary. They have some degree of reproductive isolation, as I'll say in a minute. But they're kind of, in other ways they're not reproductively isolated. 

So there are certain components that you can identify that seem to keep them apart. But they haven't gotten to a stage where everything is completely divergent yet. So we're studying the process of speciation kind of as it happens. So I like that. Another thing that's neat about mice, largely because of their commensalism with humans, is they hybridize in nature quite a bit. So the species have ranges that overlap, as I'll mention in a second, and they form hybrids. And those hybrids turn out to have problems. But it's kind of neat that we can study, we can hybridize mice in the lab under controlled conditions, and we can also look at hybrids in nature. That's helpful. And then, of course, since I'm in a genetics department and we're looking for the genes, ultimately it would be nice to do it in something that has genetic tools and the genetic tools for house mice, house mice are the premier genetic model organism for mammals, at least. 

And in some ways for vertebrates this is true. And I'm also drawn to them, to be totally honest, because of the hope that something that I understand here will tell us something about our own origins. Now, again, we haven't started doing the crosses between mice and humans because there's been some issues with getting the policies through. 

(laughter)

 But the idea is that mice have, even though they are divergent from humans by quite a while, they have mostly the same genes that we do and they are an experimental organize where we can actually manipulate things. Yeah?

- What is the picture?

- Oh, sorry. This is a very special mutant, no. I should have said that. 

So this is a black 6 mouse which is just a very commonly used biomedical strain. And somebody just drew on this thing to show this is supposed to be DNA. And this is from a project where people were sequencing the genome of house mice. So it's supposed to be some kind of clever fusion of that. So now, in the last 10 or 15 minutes, whatever, I'm just going to talk about two, quickly, two efforts to find the genes, to find the mutations that cause reproductive isolation between species of mice. So this is a sort of old map. House mice seem to have originated in this part of the world, sort of around India and Pakistan, and now there are principally three species, domesticus, musculus and castaneus. These other ones are sort of uncertain so there's basically three species. And if you find a house mouse in Madison, it's going to be Mus domesticus, almost certainly. I say "if" because we've actually been trying to trap house mice around here, and I don't know, maybe you guys can tell me about this, it seems there are a lot of deer mice which sometimes look sort of similar but are actually really divergent from house mice. 

These species we think, based on comparisons of DNA and the fossil record, diverged about half a million years ago. So that might sound like a lot but actually they're just in the process of really getting going. So one thing I want to point out is that there is this interesting area that stretches across central Europe where two of these species meet and hybridize. It's called a hybrid zone. And so now I'm just going to mention quickly what we can do with that. This is actually work that I did a few years ago. And the idea, I'll explain this in a second, the idea is that when you look at these hybrids in the hybrid zone, you can see which mutations seem to be removed from the population. So one way to say it is, if I go look at hybrids, or I sample individuals from the hybrid zone and they look like one species or the other instead of looking like a mixture and that happens consistently for a particular gene, we might suspect, if we know that they're really hybridizing, that that gene is causing some kind of problem in hybrids. Does that make sense? So we can look for, thanks to genomics and other advances, we can actually just look for those genes. 

So what I did was something pretty simple. We had these big samples of mice from the hybrid zone and got their DNA, and I just measured the rate of genetic exchange for different pieces of the X chromosome. Since then people have looked at other chromosomes, but we thought the X would be important. So what you can see here, this is a relatively high rate of exchange and this is a really low rate. I don't have the units here but this is sort of close to zero. So we know that these animals are hybrids because you can look at different parts of their DNA and see signs of that, see contributions from both parents, both species. But in this region you almost never see, in this region of the chromosome, you almost never see evidence of hybridization, despite the fact that the same animals are hybrid in other parts of their DNA. So this suggests to us that this part of the X chromosome contains genes that cause problems in hybrids. So that's a very quick summary of along many years of work. Yeah?

- How do you distinguish that from a fourth species?

- Repeat the question.

- Payseur: He asked, how do we distinguish that from a fourth species. The way that this is done is by identifying markers that are diagnostic for the two species. So we know that they differ between these two species, but there's something else happening with the other species. Does that make sense? So we can see pretty unambiguously which kind of population the hybrids came from. So that's the hybrid zone. Now I just want to spend the rest of the time talking about genetics in the lab. Since this is not Wednesday Nite @ the Hybrid Zone, it's Wednesday Nite @ the Lab. 

(laughter)

 And I'm just going to show some quick kind of patterns here. So here are our three species, and comparisons of their DNA strongly suggest that the evolutionary history of these three species looks like this. So this, for those of you who haven't seen this before, it's a phylogenetic tree that relates the different species to each other, and the idea is that the three species shared a common ancestor back here at the root of this tree and then there was separation of two lineages, one of them led to domesticus and the other one led to another common ancestor more recently between these two species. So musculus and castaneus are more closely related to each other than either is to domesticus. So because of that, we decided I didn't want to just study reproductive isolation between one species pair so we actually did it between two. So we reasoned that these two species are sort of equally divergent from this one in time. So it might be nice to compare them. So we did this cross between domesticus and musculus, and we did, using the same strain of domesticus, we did this cross between domesticus and castaneus. So we have two big crosses and just to remind you or to point out kind of how this works, the idea is that we've got, we just think about one chromosome, say it's chromosome one that we're following, and this is true for any kind of genetic mapping that people are doing, particularly in mice in this sort of scheme, and we have strain A that is marked by blue, and we have strain B that's marked by red. 

And usually what people do in this situation is to start with inbred strains, that's what we did too, so that there's no variation within A and there's no variation within B. So when I see the A version of a marker in the genome, I know it came from this parent. And when I see the B version, I know it came from the B parent. So that's very helpful. We can make these F1s just by crossing A to B. And you know that they have a copy from each parent at every piece of the genome. And then the sort of power of crosses is that we can mix these F1s with each other. And what happens is by recombination of the DNA, the different combinations, the chromosomes get all rearranged. So the individual F2 mice, some of them have configurations that look like this, some of them recover the same, they look like one of the parents for a particular chromosome, but you get a very large number of combinations. And each mouse has a completely unique genetic constitution despite all of them having come from the same parents, genetically identical parents. 

So why do you do this? The point is just that what we want to understand is hybrid sterility, which I'm going to talk about in a second, and what we want to figure out is which pieces of the chromosome are associated with hybrid sterility. So one way to do that is just to shuffle up all the different pieces and get mice with the right combinations and ask whether they're sterile. So that's what people do with genetic mappings. You can imagine each of these mice, we have some measure of sterility and we know which part of their chromosomes came from which parents and we ask whether those are correlated. Okay? And what we did, we repeated this design. So domesticus is strain A, musculus is strain B, and we did the same thing with domesticus strain A and castaneus strain B. The point is we just generate these very big crosses and measure phenotypes. So there's a lot of phenotypes that you can measure. 

We're interested in problems with the male germ line here because we knew that hybrid sterility in males was something that people had seen before in crosses between these species. The females, by the way, the hybrid females are completely fertile, as far as we can tell. But the hybrid males, if you can do the cross in one direction, that is a certain parent from one species and the other parent from the other species, then you get some issues. So we're trying to understand the genetics of those. So there's all kinds of things you can measure. And I'll just mention some of them. This slide I'm sure you've all seen before. No, I'm just kidding. 

(laughter)

 What this is, I just think it's fun to kind of look at these pictures, but what this is is a cross section through the testis of these mice after they're dead. 

(laughter)

 And what I'll just point to here, so what you see are these little tubes, these circles. And what the circles are, they're called seminiferous tubules, some of you will know this, but this is sort of where sperm get made in the testis. Which I always think is really cool to look at. So if you just take one of these seminiferous tubules, what you have is dividing cells around the rim of it and as you move inwards you're going through more and more sort of cell divisions, you're actually going through meiosis, and what happens is in the lumen, in the tube in the middle, these sperm get spit out and these eventually go and they exit the testis, go to the epididymitis and things like that. But that's spermatogenesis 101. All I want to say about this is that when you look in hybrids, you see a lot of variation in these kind of pictures but one thing that happens, you see a lot of crazy things. Like this kind of a configuration and this one, you basically have complete degeneration of the dividing cells here. So you end up not really having sperm in the tube. There's also these darkly stained pieces suggest some kind of program cell death. 

And we don't know very much about how that's happening here but it seems to be happening. I have lots of pictures like this, and you can find just about every kind of defect you can imagine. So it's actually pretty challenging to figure out how to organize them, but I won't make you do that tonight. I'm just going show, really quick, this is testis weight variation in hybrids. All I want you to see here is that the F2 individuals in each of the two crosses have a very wide range in their testis weight. There are some with very large testes, there are some with very small testes. We're interested in the ones that are over here. This is a range where we expect based on the study in mice and humans and other organisms that these animals are largely sterile. So what we want to do is figure out how these animals differ genetically from the rest of them that have kind of normal or larger testes. You can do the same thing with sperm density where we just count the number of sperm. 

And there's a very wide range of variation. And for those of you who have taken a genetics class before, this is what you look for right before you decide to try to map something. Because this variation, remember these animals are all raised in the same environment so that means that this variation is coming from genetic variation among those animals. Differences in the DNA that got shuffled. So here are some more phenotypes. You find all kinds of crazy things. I realize I didn't put what a normal sperm looks like on here so you don't know. But one thing that we see a lot is this kind of bent tail where somewhere, this is the head here, it's probably hard to see that, but the tail just has a kink in it and folds back over the head. So these sperm have trouble swimming. I don't know if you can even see this. 

You get a lot of tails without any heads. You get heads without tails. I didn't know what to call this one. So you also just get sperm heads that look really different than normal ones which seems bad. And there are, again, you have to sort of, I don't have a movie of this, but watching these sperm swim is kind of interesting. It's like they just sort of woke up in the morning, they have a cramp and they can't really go this way. Anyway, but you get all kinds of crazy defects. So this is hybrid sterility between these species of mice. Yeah?

- Looking at the F2s, the F2 generation.

- Yes.

- But the F1s are the original hybrids, right?

- Yes.

- So theoretically, if they had any of these problems, you would even be able to see it? Do you have it where the F1s don't have any offspring?

- So, that's a good point. So we can cross, right, so obviously we can cross the F1s so they're not completely sterile. If you do the cross in a direction where the musculus is the mother and the domesticus is the father, the F1 males are completely sterile. But we can do the cross in the other direction. So we end up mapping that sterility that appears in the F1s in the F2s. 

It's a lucky thing that you can do that because otherwise I wouldn't be talking about genetics here. I wouldn't be able to do the cross. That's a really good point. I'm going to try to finish up here. So one thing we can do is measure sperm head shape which has been shown to be strongly correlated with fertility. And actually most of the evidence for this is in mice and in bulls. Anyway, we can measure the sperm head shape, and what I wanted you to see here is that there are differences between the three species and their head shapes but there are also a lot of similarities. So when you look at hybrids, and this is kind of addressing your question a little bit, in one direction of the cross the sperm look kind of like the parents, there are some variation but they look sort of similar, but if you look in the other direction of F1s where the animals are sterile, you see quite a change in the head shape. Now I want to emphasize we don't know how that change really causes sterility. But it seems to be a pretty dramatic difference so we think there's probably something going on there. 

So then what we do, as I said, is we just look at markers in the genome and in the DNA, and the markers are just, as I said before, things that allow us to differentiate between the two parents and the cross. So we have red and blue and we just go through and see for every animal whether they have two copies of red, two copies of blue or one of each for every one of these locations. So each one of these is an independent marker. And you get genotype data. So this is like the rows are individual mice and then you have 200 markers in the columns and this is, like I said, sort of red/blue and green would be when you have both. There's no real pattern for you to see here, just that we have lots of variation in the markers. And now we can ask whether that variation is associated with the traits that I was talking about. So all this is, and this is I'm almost finished here, is a plot where for each chromosome we're just plotting the statistical evidence that there is an association between a trait and the genotype at each marker. So we go through and test each marker one at a time and say marker one, do you predict whether there's sterility between, can you separate sterile from fertile males? If you can, then we would say that there's linkage to that marker. 

If you can't, then it's not really useful. So when you get higher peaks here, what they mean is that there's a stronger association than you would expect just by chance. So these little asterisks are cases like that. And so we find for testis weight, for example, that there are genes on chromosome two and chromosome four and chromosome 10 that are predictive of what your testis weight is. We don't know the exact genes yet. What we know is their home on the DNA, we know their location. So we're working on trying to figure out where the genes are. You can do the same thing with sperm density, and you get a huge sign of a locus or a mutation that causes hybrid sterility on chromosome X, which is interesting, especially when you remember what I showed you about the hybrid zone. Maybe those things are related. Obviously, you can do this with all the phenotypes, which we have, and one thing that comes out over and over again is there's evidence for a locus on the X but also other changes in other parts of the genome. 

So I was trying to talk to you about the evolution of reproductive isolation. So I just described some of the genetic changes that we think are responsible for hybrid male sterility between species of house mice, but by doing the two separate crosses and knowing the evolutionary history of these three species, we can actually start to, if you will, map the history of the genetic changes that we're looking at. So let me just say that again. Imagine that you find a locus, a region of the genome that affects testis weight, for example, in the musculus cross but not in the castaneus cross. Well, our best guess of where that change happened would be on this branch because it's not seen in crosses between domesticus and castaneus. And the same thing follows the other direction. If we find a locus in the castaneus cross but not in the musculus cross, then we can map it there. And if you find changes, hybrid sterility mutations that affect both, you would think they have to happen somewhere along here because that's the, they either have to happen sort of mostly in domesticus or before musculus and castaneus split. This is kind of a minor issue but I just wanted to show you why we're doing the multiple crosses. The goal is to be able to say not just what the genetic changes are but something about where they happened and when they happened in evolutionary time. 

So I can finish up here by saying I think there are several implications of this work I just showed you, quickly. The X chromosome is important in speciation. We actually knew that before we started from work in other systems, but it looks to be true in mice as well. We've identified multiple genetic changes that contribute to speciation even in the 500,000 years since these species have been separated. This is one that, I don't have a strong claim to make about this, that's why I put a question mark, but as I said before, when you consider the fact that the same kinds of genes, we know the same kinds of genes are responsible for spermatogenesis in mice and in humans. In fact, that's the way we know what the genes that we think we know are predictive of fertility in humans, most of them were identified in mice. And then they went and looked in humans in a targeted way and noticed that the same genes were involved. So I think this is something that we can actually test. We can look in populations of humans at these genes that we find in mice and ask whether there's a relationship between the version of that gene you have and some kind of clinical phenotype, whether it be the sperm count or whatever. We haven't done that yet but I think it's probably worth considering. 

And then the larger implications I just want to impress upon you, we didn't have enough time to get into the details about this, is that despite what you might hear from more polemical kind of sources, the basic processes contributing to speciation are largely understood. This was one of the subjects that really fascinated Darwin. He didn't really figure out how it works even though the name of his book was "On the Origin of Species." But he came up with some really clever insights. And people have been studying this for quite a while. So there are things that we know well and then there are gaps in our knowledge. But the basic processes seem to be pretty well understood. Another thing I wanted to impress upon you is that, and I think this is almost always the case in biology, that when you can look at interesting problems through the lens of genetics, they become not just interesting but explainable. Genetics is one of those things where we have a mechanism, we understand something about how genes get transmitted from parents to offspring. So being able to look at phenomena like hybrid sterility in the context of genetics is a pretty powerful approach. 

And then this hopefully goes without saying. And I just want to finish by acknowledging my graduate student, Mike White, who has done all these crosses, measured all the phenotypes, and is just a terrific student to have in the lab, and I'm really blessed to have amazing students and postdocs. It's another wonderful thing about UW Madison. And then the National Science Foundation has graciously agreed to fund this research after only four applications. 

(laughter)

 Thank you, very much. 

(applause)