Jennifer Graves is Distinguished Professor at La Trobe University in Melbourne, Australia. Her groundbreaking investigations into how genomes are organised in Australia’s unique fauna have yielded insights into our understanding of evolutionary genetics, particularly the function and evolution of mammalian genomes. She is also well known for her influential studies on human sex chromosomes and sex-determining genes. Her work has won her several awards including the 2006 L’Oréal-UNESCO Laureate Fellowship for Women in Science and the 2017 Prime Minister’s Prize for Science given by the Australian Government. Graves was in IISc in March 2018 to give a public lecture organised by the Indian Academy of Sciences and participate in a discussion titled Women in Science hosted by the IISc Alumni Association. During her visit, she also spoke to Connect about how sex is determined in humans, why the human Y chromosome that specifies males might soon go extinct, and what Australia’s unusual animals can tell us about the evolution of sex chromosomes.
There are many costs of sex not least of which is the huge investment females make in producing males. But sex seems to have evolved almost as soon as eukaryotes appeared on the planet. So, why sex?
That’s a good question. An asexual female would be doing much better by cloning herself because all her genes would be passed down to her offspring instead of only half. So there must be a good reason for you to mix your genes with those of another individual. My favourite hypothesis is that the real driver for sex is genetic variation which helps you deal better with pathogens and parasites. If there is genetic variation among the members of the population, surface antigens of pathogens are going to be different in each of them. On the other hand, inbred populations with less genetic diversity are vulnerable to getting wiped out by pathogens. So that seems to be why sex, which is at least half a billion years old, is ubiquitous in plants and animals.
“My favourite hypothesis is that the real driver for sex is genetic variation which helps you deal better with pathogens and parasites”
Sex typically requires two sexes – other than in species which are hermaphrodites. In humans, we know that females have XX chromosomes and males have XY chromosomes, just as in most other mammals. How exactly does this work?
Before I answer the question, I want to remind your readers of some elementary genetics. Our DNA which is present within the nuclei of our cells is divided into smaller fragments, rolled up with proteins into chromosomes. All our genes are present on these chromosomes. We have two copies of each chromosome, one of which is from your mother and one from your father. We humans have 23 such pairs of what are known as homologous chromosomes. The chromosomes of males and females are similar except for one pair of chromosomes. These chromosomes are identical in females and are called X chromosomes. Males on the other hand have a single X chromosome accompanied by a teeny-weeny chromosome called the Y chromosome.
In an embryo if the undifferentiated gonad receives the Testis- determining Factor (TDF), it turns into a testis, which in turn makes hormones that trigger the development of other male characteristics. If it doesn’t, it becomes an ovary. TDF is the key to making male babies. Until 1990, we didn’t know which gene on the Y chromosome encodes this protein. Many people were involved in this race to identify this gene in the 1980s. The first gene that was cloned from the Y chromosome was called the ZFY. It looked like a very good candidate for sex-determination.
Now this is something that I accidentally got involved in during the 1980s. It started with a phone call from David Page [the geneticist who discovered ZFY]. He asked me if we could map this gene in kangaroos. He said that if it is the right gene then it should be on the Y chromosome in all mammals including marsupials. I gave this job to two of my students: Jamie Foster and Andrew Sinclair. And to their shock, they found that it was not on the Y chromosome in kangaroos; it was instead on chromosome 5. In another mouse-like marsupial called the dunnart, it was on chromosome 3. So ZFY was in the wrong place and so must be the wrong gene. Andrew Sinclair went to London after his PhD where he eventually cloned the right gene: SRY. We know this is the right gene because a mutation in this gene makes the embryo female even if it has a Y chromosome. And Jamie came back a few months later to my lab where he cloned a similar gene, SOX3, on the X chromosome, which is the ancestor of SRY. Then he returned to the UK and discovered SOX9, which is the target of SOX3. But the story of sex-determination is much more complicated than we initially thought with many more steps upstream and downstream to SRY.
“We know this [SRY] is the right gene [for sex-determination] because a mutation in this gene makes the embryo female even if it has a Y chromosome”
You’ve described the sex chromosomes as an example of “dumb design” to contrast it with the idea of “intelligent design” used by creationists. Could you elaborate on what you meant?
Sex chromosomes are a lot of trouble. They don’t pair very well during meiosis. They pair only at the very, very tip. That’s dangerous because it can lead to infertility. Another problem is dosage. The X chromosome has over 1500 genes and the Y chromosome only has 27 in the male-specific part [it has about 45 in all]. So females have two copies of most genes and males just one. Therefore there’s a very elaborate mechanism to silence one of the X chromosomes in females to match the males. So that could go wrong as well. Another problem for males is that there’s only one X chromosome; if a gene on it mutates, there’s no backup. So men get many sex- linked conditions like haemophilia and colour blindness. And finally, the gene that promotes sex is right at the top of the Y chromosome. And sometimes it gets accidentally recombined with the wrong chromosome and that doesn’t work very well either. So are sex chromosomes the way they are because there’s no better way to make them? My answer is no. Organisms are stuck with the genes they receive from their ancestors and they make the best of this bad situation by evolving mechanisms like making genes on the X more active to match the other chromosomes, then silencing one in females. So I think it’s a wonderful example of dumb design. You see many such examples of poor design in the human body itself. There is a tendency is to look at nature and say that there must be some good evolutionary reason for this. But sometimes it’s just a hangover from our evolutionary past.
You just said that the Y chromosome is a degraded genetic landscape. In the past you’ve referred to it as the “wimpy Y”. Why is it degenerating?
There are two things about the Y chromosome that are bad. One is that it is always in the testis by definition. It’s never in an ovary. And the testis is a very dangerous place to be because there’s a lot of cell division to make sperm. Every cell division is an opportunity for mutations. So you get a much higher rate of mutation in the genes that come from the sperm than you do with the genes that come from the eggs. And that’s true for all the father’s genes, not just sex genes. Besides, Y can’t repair itself very well unlike the other chromosomes. For instance, if you have a mutation at the top of one X and one at the bottom of the other X, when they pair, they undergo a process called recombination. So you can combine the good bits of the two X chromosomes and get a good X in the egg. The Y is all alone in the world – it doesn’t have the opportunity to recombine and get rid of the mistakes. So the lack of meiotic crossing over of genetic material and the high mutation rate mean that its genes are very vulnerable and the chromosome degenerates very fast.
You’ve also predicted that the human Y chromosome could go extinct in a few million years. Given the rate of loss of genes in the Y in the past few million years, what is the prognosis for this chromosome?
All placental mammals and marsupials have Y chromosomes. But they are all a little different. The mouse Y has lost almost everything. There are only about two genes that you need to make sperm. And in fact in some rodents the Y has completely gone, and has been replaced with something else. The primate Y has been stable for some time. But that doesn’t mean it’s going to be stable forever. It could degenerate very rapidly. It’s full of repetitive sequences; so deletions are quite common. Many of these cause infertility. If you assume that degradation is linear, you can easily work it out. We know from studying the genomes of other mammals that our XY pair started to differentiate about 166 million years ago. Originally there were 1669 genes on it, and today there are only about 45. So you can work out that about 9.8 genes disappear every one million years. At that rate, the rest of the Y chromosome is going to be gone in 4.6 million years. But of course this process is probably not linear. So it could be gone sometime between next week and in the next several million.
“At that rate, the rest of the Y chromosome is going to be gone in 4.6 million years”
When that does happen, males, and therefore the human species, could go extinct. Or a new sex-determination system could evolve…
Yes, it’s interesting to think about it. One possibility that people love to champion is that we’ll become a species with just females who reproduce by parthenogenesis. There are lizards that do it. But that wouldn’t work with humans or other mammals because of genomic imprinting. We know that there are about 30 genes that become active only if they come through the sperm. They’re very important genes for development. So we do need men, we do need sperm, and without them, we’re all going to go extinct. Unless, as you say, a new sex-determining system arises. And that’s actually happened much more frequently, and more recently, than you might think. It’s quite easy to make new sex-determining genes. There are rodent lineages which have already lost their Y chromosomes, and a new sex-determining gene has evolved very recently. If you came back in 4.6 million years, you may find that another chromosome will have become the sex chromosome in humans.
In one of your talks you seemed to suggest a speciation angle to this story.
When you look at the big picture, at every major rearrangement of mammal sex chromosomes, you see divergence of major groups of mammals. So I did begin to wonder if sex chromosome evolution drove these major separations. And I think there’s quite a bit of evidence that sometimes speciation is driven by a chromosome change, particularly a sex chromosome change. Because when it changes, you really mess up sex. And that serves as an effective barrier between two divergent populations. So I’m suggesting that once you have a sex chromosome change or if you have the evolution of a novel sex-determining gene, that’s going to drive speciation. And I think that’s what’s happened in these rodents that have lost their Y.
“Once you have a sex chromosome change or if you have the evolution of a novel sex-determining gene, that’s going to drive speciation”
Tell us more about these rodents.
It’s a group of rodents called Japanese spiny rats…I love them. There are actually three species. One of them still has a Y chromosome; it’s a bit of a strange Y, with 100 copies of a mutated SRY that doesn’t work well. But the other two species have lost their Y and I’m suggesting that it’s the evolution of a new system that drove speciation there. And exactly the same thing seems to have happened with the mole voles in eastern Europe. Again they have one species with a Y chromosome, one species with a single X chromosome in both sexes, and one species with two X chromosomes in both sexes. So again you see these species which diverged very recently have different sex-determination systems.
Let’s talk about the X chromosome now. You have used the term “brains and balls genes” to describe the genes on X. What do you mean by that?
The X and Y chromosomes are similar only at the very top – that’s how they stick together during meiosis when the sperm is made. The rest of the X chromosome is very different, even when compared to other chromosomes. There’s been an accumulation of way too many genes that have to do with both reproduction and brain function on the X chromosome. And many of same genes are active in the brain and the testis, and code for both reproductive and cognitive traits. So when there’s a mutation in them, it leads to sex-linked intellectual disabilities which are also accompanied by abnormalities in the gonads or infertility. And this is seen mostly in males because they have only one X. These are what I call the “brains and balls” genes; the term was actually coined by my German collaborator Horst Hameister. But why has this happened? The brain and the gonads are of course very different organs. The best explanation I can think of is that different types of selection have acted upon the same large multifunctional proteins, giving them functions related to intelligence and reproduction. If a mutation confers an advantage to males, it’ll be immediately selected because, as I just mentioned, males have only one copy of genes on the X chromosome. And I like the theory that intelligence in males is selected by females who are looking for smart partners! I’m waiting for somebody to look at what these proteins actually do and how they bind in the testes and the brain – they could have different binding partners.
“There’s been an accumulation of way too many genes that have to do with both reproduction and brain function on the X chromosome”
Your main research is in the field of comparative genomics. What do the genomes of marsupial and egg-laying mammals, which are native to Australia, tell us about the evolution of sex chromosomes in mammals?
What we see in marsupials is the same kind of sex-determination system with an X and a Y. Again, the SRY on the Y looks like the sex-determining gene. And it turns out that they are the original X and Y of therian mammals [marsupials and placentals]. In early placental mammals, a bit of an autosome [non-sex chromosome] got stuck on to it. The human Y is practically all derived from this recently added bit. We know that in the elephant X, for instance, the bottom part is old and the top part is new. And it’s almost the same as in humans. The only difference between human and elephant chromosomes is the centromere [place where chromosomes attach during cell division] has moved slightly. So what we can tell by studying marsupial and other mammals is that mammal sex chromosomes evolved rather recently. But the real key to understanding the origin of sex chromosomes in mammals is the platypus. These are bizarre egg-laying animals. Platypuses and all other mammals had a common ancestor about 165 mya [million years ago]. They are mammals of course – they have fur and feed their young with milk, but retain many reptilian characteristics – but their skeleton is more like that of a lizard and the male makes venom much like snake’s venom. They also lay leathery eggs like those of snakes. And their chromosomes too are very different compared to those of other mammals. They do have the old X and the bit that was added later on, but these are on autosomes. So they have completely different sex chromosomes with no homology to human or even kangaroo sex chromosomes. Remarkably, they have a lot of homology to bird sex chromosomes! We think what they have is probably what the first mammals had, but then it changed into the new XY sex-determination system just before the evolution of the marsupials about 150 mya.
“The real key to understanding the origin of sex chromosomes in mammals is the platypus”
So how is sex determined in platypuses?
It is really weird because they have ten sex chromosomes. They have five Xs – with two copies of each in females and one in males. And there are five Y chromosomes that are male specific. We used a technique called chromosome painting to mark the Xs and Ys and showed that each sperm has only Xs, or only Ys. During meiosis we see these ten chromosomes all lined up and we think all the Xs go to one pole and the Ys to another. So you get two kinds of sperm: one having five Xs and the other having five Ys. It’s a crazy way to do sex. But it works quite well. And there’s no SRY gene. We think another gene, AMH – the anti-Mullerian hormone gene – which we find on one of the Y chromosomes, is the sex-determining gene.
Reptiles are also fascinating because in some species sex is determined by the temperature of incubation of the eggs. But you have found something even cooler in the Australian dragon lizards…
The Australian dragon lizard – a beautiful little creature – has genetic sex-determination, but a number of its closely related species have temperature sex-determination. We identified sex chromosomes and got the sequence from them. In fact, we sequenced the entire genome of this particular species a few years ago and we think we know the sex-determining gene. While studying this lizard, we found something really strange. At its usual range of temperatures, sex is determined by their chromosomes. Boys have ZZ chromosomes and girls have ZW chromosomes. But when we incubated the eggs at a higher temperature, they were all girls. We already had molecular markers for their sex chromosomes by then. We were able to show that we had both normal ZW females and also sex-reversed females with two Z chromosomes like males. What’s more is that they were viable and fertile; in fact ZZ females do better than the normal ZW females – they lay more eggs and their hatch rate is better. The nice thing is that we could mate ZZ females with ZZ males. All of their hatchlings were ZZ, and their sex is completely temperature-dependent. So we were able to switch a sex-determining system from genetic to environmental in just one generation. The scary thing is that this is actually happening in the wild because of increasing temperatures due to climate change. We’ve been sampling for ten years and during that period, there’s been a big increase in the numbers of sex-reversed females. So at this rate they’ll all be females in next few decades. So you have to worry about climate change not just for species with temperature- dependent sex, but also those with this over-ride system like this dragon lizard. It might be more common than we think – there’s a skink in which cold temperatures induce sex-reversal in males. So obviously a lot of things can happen with extremes of temperatures even when you’ve got sex chromosomes.
I’d also like to add one thing here. We’ve been able to use the dragon lizard to understand how TSD [temperature sex- determination] works. We looked at the transcriptome (the RNA made from active genes) of the ZZ sex-reversed females and found some really spectacular changes. We found that the stress genes were elevated; so it looked like high temperatures were inducing the stress pathway. That’s not such a surprise because we think stress is involved in sex-reversal in fish as well. But we also found that two genes had a really unique transcript that could not code for the normal protein. These genes are big players in epigenetic silencing. So we now have the first clues of how the environment affects the genome.
“The scary thing is that this is actually happening in the wild because of increasing temperatures due to climate change”
In the next few years what are the big-ticket questions in this field of sex-determination that you would like to see answered?
I would love to understand how the sex-determination systems change. We have many examples where they have very recently changed and turned over. Even better, in an experimental system, we actually might be able to build a sex chromosome, and see what happens in real time in the lab. And that’s not an impossible dream. As I mentioned already, the sex-determining pathway is very complex. We thought it would be simple. We now know of at least 30 genes in this pathway – some promote testes and some ovaries, some antagonise testes and some ovaries. This is again an example of dumb design because these pathways get built up and changed all the time – sex is so diverse. Mapping this process is going to be important. I would also like to see how the environment interacts with these pathways. We have the genomes of many species sequenced. We don’t lack the data anymore; we have to know where to look.
You study a subject that can be controversial to some people. Have you received any pushback, either from the extreme right or left, especially given how politically charged our world is right now?
It’s never been an issue in Australia. It’s been an issue for some other people in the same field. And they tend to be men. I think that’s worked in my favour for once. Not many things work in favour of a female scientist, but I think I have been able to say things that would be hard for a man to say. But a few years ago, I started to get strange invitations from some feminist magazines with words like “hermeneutics”, the meaning of which I didn’t even know. And I realised that they were using my work to further a political agenda. I thought that was rather unfortunate. It is not political in the traditional sense, but it does feed into all sorts of social tensions of gender and gender politics. In some quarters I am unpopular because I cannot pretend that women are genetically identical to men (see my article “Not just about sex: throughout our bodies, thousands of genes act differently in men and women” in The Conversation).
“In some quarters I am unpopular because I cannot pretend that women are genetically identical to men”