02 March 2017

A mystery gene in human brain development: discovering ARHGAP11b

The human brain is often described using literally cosmic superlatives. Here is V.S. Ramachandran, a renowned neuroscientist:

The human brain, it has been said, is the most complexly organised structure in the universe and to appreciate this you just have to look at some numbers. The brain is made up of one hundred billion nerve cells or "neurons" which is the basic structural and functional units of the nervous system. Each neuron makes something like a thousand to ten thousand contacts with other neurons and these points of contact are called synapses where exchange of information occurs. And based on this information, someone has calculated that the number of possible permutations and combinations of brain activity, in other words the numbers of brain states, exceeds the number of elementary particles in the known universe.
That's some serious complexity there, yes, but hidden in the description is a mundane reality: the human brain is made of neurons which make synapses with other neurons, which means that it's made of the same stuff as the brain of a sloth or a goldfish or an earthworm. (Or even a parasitic mite on an earthworm.) Still, whether or not the human brain is really special or just big, something has caused it to grow in ways that differ from its predecessors. Especially at key junctures in the process, human brain development must depart from boring old mammalian brain development. And this should be reflected by — and perhaps explained by — patterns of gene expression.

Wieland Huttner's group (at the Max Planck Institute in Dresden) has been studying brain development for a couple of decades. Recently, they did a straightforward but fruitful experiment designed to detect human-specific patterns of gene expression during brain development. They looked specifically at genes that might underlie the expansion of the cerebral cortex in the human brain. The paper is "Human-specific gene ARHGAP11B promotes basal progenitor amplification and neocortex expansion," by Florio et al., published in Science in early 2015.

It's worth taking a moment to consider the human cerebral cortex. The cortex is the wrinkled stuff that covers nearly the whole brain. A glance at the human brain can give the impression that it's a big bowl of wrinkled gelatinous glop. But that wrinkly stuff is really just at the surface. A deeper look presents a weirder story, in which the wrinkly stuff has oozed out over the surface of the brain, growing out of control until the whole outside of the brain is wrinkly cerebral cortex.

That's pretty much what happened, specifically in mammals. Look at the picture on the right (click to enlarge). The wide range in brain size is obvious at a glance. If you look more closely, you can see that the wrinkly stuff dominates in many mammals, masking the fact that the basic organizational plan of a tetrapod brain is unchanged. The elements of the cortex are very old, surely older than tetrapods. But the expansion of the cortex is largely a mammalian thing. And the expansion in humans is very striking, to the point that at least 75% of the adult human brain is cortex.

So the mammalian cortex is big. But the human cortex is, well, super big. And we'd like to understand how that happens during human development. Well again, in some sense it's just the same old tissue, made of the same old cells, as a tiny fish brain. So maybe the main difference between a big brain and a little brain is the number of times that the relevant cells in the embryo underwent cell division. If you think about it, that could be sufficient to explain how any tissue in any animal gets to its final size. This is an approximation, but if every cell in a particular organ divides once, the organ doubles in size. And since tissues and organs are built by generations of cells, then it follows that increased division of early-generation cells can make a significant difference in how big a tissue or organ can become. Here's what I mean: if a single founder cell divides once, and then each cell in each subsequent generation divides once, then after 10 cell generations we have 2^^10 cells, which is about 1000. You can double that by simply using two founder cells instead of one. And so on. The point is that the awesome, massive human cerebral cortex owes its existence, at least in part, to the behavior of the founder cells. Somewhere along the line, probably early in the process of brain development, something changed about those founders. Either there are more of them, or they are more prolific in their division, or both.

So, back to the work by the Huttner group. They decided to compare the brain founder cells of humans to the same cells in mice. As far as we can tell, humans and mice build the cerebral cortex in the same basic fashion: some founder cells are established, then these cells divide to make subsequent generations of cells that also divide, then other really interesting things happen, and then you have this multi-layered computational marvel called the cerebral cortex. The mouse cortex is less obviously layered and of course it's far smaller, but otherwise it seems the same. A comparison, then, could reveal how the human process differs.

Comparisons like this have been done before, and have revealed some influences that can begin to explain the big mammalian cortex. But it's harder to do the comparison directly to humans, and the previous work had never looked directly at some of the most important founder cells. These are called neural progenitor cells, or NPCs. I will just call them founder cells.

So Huttner and colleagues started by developing a way to get a pure sample of founder cells from the relevant stage of development. Using a combination of fluorescent dye probes and a technology called cell sorting, they winnowed the founder cells out from among all the other cell types in the developing brain, to the point that they not only could collect founder cells in pure form, but even could subdivide them by type. Specifically, they collected a kind of founder cell called basal radial glial (bRG) cells. These cells are known from years of study to be important for cortex expansion, but no one had ever gotten human bRGs alone in a test tube. This is the technical feat achieved by the Huttner group, and was independently accomplished by another group (Chris Walsh and colleagues at Boston Children's Hospital and Harvard Medical School) at the same time (both papers were published at about the same time in 2015).

Once they had the special human founder cells purified, they could compare them with the same cells from mice to ask that straightforward question: what, if anything, is different or special about human founder cells in the cerebral cortex? This is done by identifying all of the genes that are turned on in those cells, and that's why they needed to have a pure sample of just those cell types.

First, they described a set of genes that aren't human-specific but are turned way up in the human founder cells. The results of that analysis are interesting but not mind-blowing: they found genes involved in cell-surface processes that were known to be a little different in human cortex. And more to the point, these are genes that are present in mice (and thus probably in all mammals). So, yes, these are interesting clues to how the human cortex is built. Evolutionarily speaking, however, the genes aren't new or unique.

But then Huttner and colleagues looked for human-specific genes among the ones activated in the human founder cells. I know I've mentioned this a few times already, but I think it is remarkable that they found human-specific genes at all. In fact, there were 56 genes that were both human-specific (compared to mouse) and turned on in founder cells more than in other cell types. Because they wanted genes that were truly specific to the founder cells and not to any other cell in the brain, they set some strict criteria that pointed to exactly one human-specific gene that was turned on strongly and specifically in the human founder cells. That gene was ARHGAP11b, a gene that had not been described before, in any experiment on any cell or tissue.

That's how ARHGAP11b was discovered — in a deliberate search for genes that are turned way up in the founder cells that generate the cosmically huge human cerebral cortex. The scientists' success relied on two main advances: their own technical achievement of getting pure samples of human founder cells, and the existence of technologies that allowed them to identify and measure the expression of every single gene in those cells. That technology is called RNA-seq, and it didn't even exist until 2008.

I still haven't told you what ARHGAP11b does. Patience!


Image credits:
Four primate brains, Figure 2 from Hofman (2014) Evolution of the human brain: when bigger is better, Frontiers in Neuroanatomy.
Mammalian brain assortment, Figure 3 from Herculano-Houzel (2009) The human brain in numbers: a linearly scaled-up primate brainFrontiers in Human Neuroscience.
Labeled founder cells in developing mouse cortex (notice bRGs), Figure 1C from Florio et al., linked and cited above.

20 February 2017

When a GAP is not a GAP: ARHGAP11B, the mysterious human-specific gene

A truly human-specific gene, not merely a human-specific version of an animal gene or a mammal gene or a primate gene — that is something particularly interesting. Given that the human genome is 96% identical to that of our closest relatives (chimps and bonobos), and given that so much of those genomes is composed of mobile elements that are unlikely to end up being genes at all, I and perhaps others long thought that human-specific genes would be something pretty rare.

But there they are — genes by every definition, that code for protein and are expressed in human tissues, that are unique in humans. One of the most interesting is a gene that brings together some of my personal favorite topics in biology: brain development, cellular signaling systems, and of course evolution. The gene goes by the unfortunate "name" of ARHGAP11B.

I do consider ARHGAP11B to be a unique human gene, but its name betrays its evolutionary history and its membership in a family of genes, so it's not completely unique (specifics to come). That family is the family of GAPs, a group of proteins that were the focus of my postdoctoral research years ago. GAP stands for "GTPase-activating protein," and besides being a typical morsel of biochemical jargon, the phrase is a bit of an insult to the roles played by these proteins in cellular signaling systems.

A RhoGAP domain
The signaling system of interest here involves a superfamily of proteins called small G-proteins, or small GTPases. The 'small' refers to their actual size, and the rest describes a biochemical detail of their function. Both aspects of the name are a hindrance. Because while these proteins are indeed small, their roles in multiple biological processes are huge. And while they do in fact engage in the biochemical process of hydrolyzing GTP (it doesn't matter whether you understand that at all), this is not what they do. What they do is throw switches to activate major changes and activities within cells. When the small GTPase is tagged with GTP (again, the details are unimportant), it is on. And when the GTP is cleaved to GDP, it is off. To turn the switch on, the cell has to stick GTP onto the protein. This is done by proteins called GEFs (that stands for guanine nucleotide exchange factors). And when the cell wants the switch off, it uses GAPs. The GAP activates the GTPase, which cleaves the GTP to GDP.

The basic overview of that is: small GTPases are switches that are major players in signal systems inside cells. GEFs turn them on. GAPs turn them OFF.

It's hard to think of a basic cellular activity that isn't controlled by these switches. One famous system is the so-called MAP kinase pathway. This pathway is used in lots of contexts to do lots of things, but one common use is to stimulate cell division. (For this reason, chemicals targeting various players in that pathway are candidates for anti-cancer drugs.) One of the key focal points of the MAP kinase pathway is Ras, and it's possible that you have heard of this famous protein. It's a small GTPase, and in fact the superfamily of small GTPases is called the "Ras superfamily" in its honor. When the Ras switch gets stuck in the on position, the typical result is uncontrolled cell division. This means that Ras is a common oncogene, and whole families of cancer are defined in part by the presence of mutant Ras that is stuck in the on position.

One subfamily of small GTPases takes its name from the first known member: Rho. The members of this subfamily specialize in a set of cellular functions that almost always involve the cell's skeleton (aka the cytoskeleton). This means that these proteins are centrally involved in how cells acquire their distinctive shapes, how big or small they get, and notably how (or if) they move through the body.

This brings us to ARHGAP11B. This human-specific gene encodes a protein that looks a lot like a RhoGAP. It is basically identical to one part of another gene, found throughout the animal kingdom, called ARHGAP11A. That gene serves as a GAP for Rho proteins, and this means that it flips the switch on these important regulators to off. So it would be reasonable to hypothesize that this is what ARHGAP11B does.

But on the other hand, ARHGAP11B is a drastically shortened version of ARHGAP11A. ARHGAP11B is made of a piece of the business end of ARHGAP11A, with a string of other material stuck onto the end. That "string of other material" is a snippet of protein sequence, a string of 47 amino acids (green in the diagram below), that is not found in any other genome. Not in the mouse, not in chimps or bonobos or orangs. The protein made by the ARHGAP11B gene is truly unique to humans (including Neanderthals and Denisovans). Nevertheless, it starts off with about 230 amino acids that match the business end of ARHGAP11A, the part that makes ARHGAP11A a RhoGAP, the part that makes the protein an off switch for Rho proteins. That part, called the GAP domain, is about 250 amino acids long (it's in purple in the diagram below), and ARHGAP11B is only missing the last 26 amino acids of this domain. So still, it would be reasonable to hypothesize that ARHGAP11B functions as a RhoGAP.

Now, I haven't told you the whole story. The way the discovery happened has next to nothing to do with GAPs and domains and biochemical reactions. The scientists who discovered this new human gene were specifically searching for genes that were uniquely expressed in the developing human brain. The gene that really jumped out in their experiment was ARHGAP11B. So they knew that the gene was potentially important at a key stage of brain development. I mention this because once they saw that the gene was related to a RhoGAP, and that it contained a nearly-complete GAP domain, I suspect that they hypothesized that the gene encoded a RhoGAP.

So they tested the hypothesis. And the results explain why I think the story of ARHGAP11B is truly remarkable. The protein does in fact exert an important influence on brain development, which I will discuss in the next post. But it has no RhoGAP activity. The loss of those 26 amino acids from the GAP domain results in the erasure of the biochemical activity that the original gene (ARHGAP11A) always had.

And yeah, that means I made you read a bunch of stuff about RhoGAPs that ended up being irrelevant. But RhoGAPs are cool, so tough. And more seriously, because we don't yet know what ARHGAP11B does accomplish at the biochemical level, it remains possible that Rho proteins are somehow involved.

Why is this case so remarkable? It's this:

The protein encoded by the ARHGAP11B gene is a unique human gene with a new and thus far unknown biochemical function. It clearly arose from a RhoGAP gene, but the protein is not a RhoGAP. It's something else. Something new. And whatever the protein is doing at the level of biochemistry, the result is increased growth of a specific part of the human brain. That's for the next post.


Image credits: RhoGAP domain from the Structural Genomics Consortium, ARHGAP11 diagram from Figure 1D, Florio et al., Science Advances (2016).

23 January 2017

Does it take special genes to make a special human?

We humans think we're pretty special. Here's Hamlet, in the speech that gave this blog its name:

What a piece of work is a man! How noble in reason! how infinite in faculty! in form, in moving, how express and admirable! in action how like an angel! in apprehension how like a god! the beauty of the world! the paragon of animals!

Hamlet, Act II, Scene II, The Oxford Shakespeare
It is common, at least in the West, to consider humans "the paragon of animals," typically making reference to the ancient idea of "God's image." I won't address here whether humanity is a paragon, but here's a more tractable question: what are the facets that distinguish humans from other animals? Biologically speaking, what is special about humans that sets them apart from other apes?

Anatomically, we're pretty unremarkable apes. We have enlarged gluteal muscles and other adaptations that make us good distance runners. We have fancy thumbs that make us good tool users. We have a tweaked larynx that facilitates speech. We have spineless penises that facilitate other things. And of course we have what Hamlet was talking about: big brains and associated cognitive abilities.

These biological specializations do make us human, and that makes them interesting and important. But biologically, I have always thought of them — all of them — as incremental changes to an ape blueprint. I have never thought of human biology as something extraordinary. And for that reason, I have always been skeptical of attempts to find human-specific genes or proteins or cell types that would explain human specialness.

Now, I don't mean that I don't think there are human-specific changes in the genome. Every species has species-specific genetic components, essentially by definition. But I never expected that we would find extraordinary new genetic components in the human genome, since I never thought our biology was extraordinary. Same goes for special cell types: some really smart biologists have looked for human-specific nerve cells in the brain. The human brain is big and complex, but it has always seemed to me to be built of the same stuff, in the same basic ways, as all the other brains I studied in my career.

Here is an example that illustrates my point. FoxP2 is a control protein involved in brain development, and because mutations in the FoxP2 gene lead to loss of language in humans, its roles and evolution are of intense interest. It was first thought perhaps to be a human-specific gene, indeed perhaps "the language gene," given its strong link to such a special human trait. But no: FoxP2 is an ancient gene found throughout the animal kingdom. The human version is different from the chimp version by just two amino acids. This is pretty much the opposite of a "human-specific gene." It's an animal-specific gene with some tiny human-specific edits.

But lately we've been learning that there are a lot of human-specific genes that are not so easily dismissed. The ongoing annotation of the human genome has revealed a somewhat surprising number of genes that are not found in other apes or mammals. Ed Yong has nicely explained why many of these genes were missed for so many years. And he highlights two of the more striking examples of human-specific genes: HYDIN2 and ARHGAP11b. (I'm so very sorry about our naming conventions. I wasn't consulted.) While both of these new genes are known to have arisen from copies of existing genes, both have been modified significantly. So they both seem (to me) to be "new genes." And, remarkably, they are both centrally involved in human brain development.

That last part is an especially interesting twist, since brain overgrowth is one of the most attention-getting human specializations. I think it raises the possibility that the human brain is not just a big ape brain, but a peculiar ape brain. In other words, if we found some weird new genes in the human genome and showed that they do weird new things to brain development, I might start to take more seriously the suggestion that the human brain is a distinctive evolutionary innovation.

I know that these dichotomies (merely big vs. peculiar, different vs. distinctive) are not very well defined. But I hope the basic questions seem interesting to you:
1) Are there unique human genes that underlie human specializations? Specifically is ARHGAP11b an example of this?
2) If so, what do these genes do and how did they arise in evolution?

Ed Yong has told the HYDIN2 story so far. But what about this ARHGAP11b? We now know how this gene acts on the brain and — this is really cool — we know how the gene was born. Both are up in the next post.


Image credits:
Top, Kemble as Hamlet, Wellcome Images
Bottom, Variation of brain size and external topography, Figure 7 from "The evolution of the brain, the human nature of cortical circuits, and intellectual creativity" by J. DeFelipe, Frontiers in Neuroanatomy, 2011.



05 January 2017

Relaunch in 10...9...8...

Quintessence of Dust has been on hiatus for more than five years. It's time to resurrect it. Why now? Because it's 2017, and 2017 is not a time to be quiet.

The first project involves some remodeling. Quintessence of Dust was built almost ten years ago, with a set of themes and goals that don't all fit in 2017. Most notably, the blog was conceived when I was a Christian, and for five years addressed issues and questions that I knew to be of interest to evangelical Christians. I am happily no longer a Christian, and will remodel the blog to reflect that. I do still live in the United States, in 2017, where evangelical Christianity exerts significant influence. And I know a lot about that world. So religion will be an occasional, if tangential, topic. But now I will write as a skeptic, as one who has transitioned from Christian humanism to just plain humanism. The remodeling of the site is mostly to make this clear. I do think I'll keep the Celtic cross in the banner.

In parallel with the remodeling I'll start writing about cool science. And I've already found the topic of my first post or two: a paper from last month that identifies a single mutation in the human genome that may explain (at least in part) the dramatic expansion of the cerebral cortex that occurred in our lineage. The story is a remarkable confluence of topics very dear to me: evolution, developmental neurobiology, and cellular signaling systems. The protein at the center of the story is closely related to the proteins that I spent my postdoctoral fellowship trying to understand. I'll explain all of this in the posts to come.

If you want to have a peek at the story, check out the news piece at the BBC, or the new paper itself (it's open access). The first part of the saga, in which the protein's role in brain development was discovered, was published in 2015 (also open access but requires free registration).

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Image: By Internet Archive Book Images [No restrictions], via Wikimedia Commons

16 December 2014

December 2014 update: Quintessence of Dust is on a long-term hiatus, but I haven't closed it down just yet. I have some new writing projects in various stages of planning, in collaboration with some old friends. See my About page for information about me and the blog, including current contact information.

28 September 2011

If it's not natural selection, then it must be...

The folks at the Discovery Institute (DI) are engaged in an extensive attempt to rebut my friend Dennis Venema's critiques of Stephen Meyer's surprisingly lame ID manifesto, Signature in the Cell. There are several aspects of this conversation that I hope to address in the coming days and weeks, but one jumped out at me today: the consistent confusion about natural selection in depictions of evolutionary theory by design advocates.

Consider this excerpt from a recent blog post by a writer at the Discovery Institute:

...we need a brief primer in fundamental evolutionary theory. Natural selection preserves randomly arising variations only if those variations cause functional differences affecting reproductive output.
A few sentences later, the same claim is repeated:
Indeed, given that natural selection favors only functionally advantageous variations, ...
Those claims were first made in a piece written by unnamed DI "fellows" mocking the work and conclusions of Joe Thornton, an evolutionary biologist at the University of Oregon and the University of Chicago. And the claims are badly misleading.

26 September 2011

Common ancestry, bottlenecks, and human evolution

Human evolution has been in the news quite a lot recently.

  • New genetic data suggest that ancient humans included both Neanderthals and Denisovans, which colonized different parts of the world but subsequently interbred with so-called modern humans and left telltale traces of this history in the genomes of living humans.
  • New analysis of current genetic diversity suggests that human population size underwent interesting fluctuations throughout the history of our species, but concludes that the population never dipped below a few thousand reproducing individuals.
Unsurprisingly, these findings have been discussed in the context of Christian views of human origins. In the context of some of these discussions (among Catholics, for example), I have noticed some confusion regarding the implications of common ancestry. I will illustrate the error with a stylized example, then explain why it is an error.

23 September 2011

Harmful genes, and sneaky, too: Genetic hitchhiking in the human genome

ResearchBlogging.orgGenetic hitchhiking is thought to be an inevitable result of strong positive selection in a population. The basic idea is that if a particular gene is strongly selected for (as opposed to selected against), then the chunk of the genome that carries that gene will become very common in the population. The result is a local loss of genetic diversity: all (or nearly all) of the individuals in the population will have that same chunk of genetic information, whereas before the selection process acted, there might have been a lot of variation in that chunk throughout the population. And this means that areas of the human genome that are less variable between people are suspected sites of recent positive selection. Within that chunk, there are potentially many genes and genetic elements that became more common in the population by virtue of their placement near the gene that was actually selected for. Those other genes are the hitchhikers. And it's likely that some hitchhikers are bad news – they're harmful mutations that would normally become rare or extinct in the population, but instead have become common by hitchhiking.

In the last few years, large amounts of genetic information have become available that have enabled biologists to look for evidence of such phenomena in the human genome. Specifically, two major projects have collected genetic data for the purpose of analyzing genetic variation among humans. One project, the International HapMap Project, mapped and quantified sites in the human genome that are known to vary among humans by a single genetic letter. These sites are called single nucleotide polymorphisms, or SNPs (pronounced "snips"). The project has mapped millions of these sites in a group of 270 humans representing various lineages. Another project that has made the news recently is the 1000 Genomes Project, which also seeks to provide a picture of human genetic variation using more people (more than 1000 at present) and slightly different technology. Efforts like these have taken analysis of the human genome to a new level. No longer do we merely wonder what "the" human genome is like – we can begin to learn about how genetic differences give rise to biological differences such as susceptibility to particular diseases.

22 September 2011

New limbs from old fins, part 3

The third installment of my series at BioLogos is now up. It discusses the developmental mechanisms that underlie the construction of limbs, and the striking fact that these mechanisms are the same ones used to construct fish fins. Watch for an appearance by Sonic Hedgehog.


19 September 2011

Genetic hitchhiking in English

The next post will discuss recent evidence for genetic hitchhiking in humans. So, what do we mean when we say that genes can hitchhike? To make sense of this phenomenon, we first need to review chromosomes and sexual reproduction.

Most people know that sexual reproduction creates offspring that are genetically distinct from both of the their parents. That's true, but the genetic scrambling that occurs is more significant than is sometimes reported. Let's start by looking at chromosomes.

Like every other animal (or plant or pretty much any other organism), your genetic endowment is carried in chunks of DNA called chromosomes. You have 23 of these chunks, which are rather like volumes in a set of encyclopedias. More completely, you have 23 pairs of these volumes; one set was contributed by your mother and the other by your father. Each of your parents had a complete set, also consisting of a set from Mom and a set from Dad. When your mother made the egg that became the zygote that became you, she provided you with one copy of each volume in the set, and she chose those copies randomly. For example, she may have chosen her dad's copy of chromosome 1, but her mom's copy of chromosome 2. Just by virtue of this random picking process, she made an egg with a shuffled version of her own genetic cards. Dad did the same when he made his sperm, and so your genetic complement is an amalgamation of your parents' genomes which were amalgamations of your grandparents' genomes, and so on.