pigmentation

I really can't get over how much work went into the cichlid pigmentation paper that's out in the current Science (Roberts et al. 2009). The paper examines the genetic basis for a "orange blotched (OB)" phenotype. It's a simple kind of genetic question, and it hooks into a story that evolutionary biologists like -- sex conflict in gene expression.

But wow how much work they did to iron out all the details. An example:

Pedigreed families from several species of the genera Labeotropheus and Metriaclima were genotyped for newly identified microsatellite markers on LG5 (46 families, 678 individuals, 349 OB individuals; table S1). Breakpoint analysis localized OB to an interval of less than 1 cM, corresponding to a region from 3.9 to 4.0 Mb on Tetraodon chromosome 11 (Fig. 2A). We then used association mapping in natural populations to pinpoint the causative locus. Single-nucleotide polymorphism (SNP) markers were developed spanning the region, including an intronic SNP marker for each annotated gene within the OB interval. We measured linkage disequilibrium (LD) between OB and SNPs within natural populations of Labeotropheus, Metriaclima, and Tropheops from the northern and southern parts of Lake Malawi (Fig. 2B). Within each population, a peak of LD was found in a region overlapping the interval defined by breakpoint analysis (Fig. 2, C to G). Given the shared pattern of association across study populations, we created a lake-wide mapping panel to increase the effective size of the mapping population. This lake-wide panel contains BB and OB individuals from 36 distinct populations of 12 species segregating OB (table S2). Analysis of marker data across these populations increased the resolution and statistical significance of LD beyond that available from any single population (Fig. 2H) (taxonomic names not italicized, because I'm in a Viennese cafe and too lazy right now, OK?).

This is the kind of stuff that a few years ago took multinational networks of cooperation; now it can be done in the context of a single lab with association mapping assisted by field biology.

It helps a lot that the trait is conspicuous and basically Mendelian. I would guess they had a strong presumption that it would be near the sex-determination locus, since the OB phenotype is very rare in males. They even end up concluding that the rare males who have it are probably products of some alternative sex-determining mechanism. So there were hints -- and the hints went along with the idea of testing the hypothesis of sex conflict.

But you can see the potential here for widespread genomic work on cichlids. There are an endless number of questions, with hundreds of species in the three big African lakes. These are Lake Malawi cichlids in this paper (for the most part), but Victoria cichlids in particular are so young that you can figure most of their phenotypic differences will be genetically simple -- it's like postglacial stickleback evolution, but with hundreds of different switches. And each one is telling a story about the segregation of original genetic variations in rapidly speciating populations, the spread of new alleles under rapid selection, and the tectonics of the adaptive landscape that occur as genetic backgrounds quickly shift.

All this rich field biology, with the bonus that you can keep a bunch of inbred lines in fish tanks to test for Mendelian ratios and other classical genetic observations (as they did in this study).

Although Lake Victoria cichlids haven't been around very long, their generations are a lot shorter than ours. So if you were thinking to compare the cichlid case with human timescales, like since the Last Glacial Maximum, think of what those fishes may have been like 1000 years after they reached the lake. They probably speciated fast, and we (of course) haven't done so at all. Yet in other respects this may be an interesting comparison -- and may be even more so as biologists move beyond the conspicuous pigmentation (and we of course do have many recent pigment variations...) and consider the genetics of habitat specializations and diet adaptations.

References:

Roberts RB, Ser JR, Kocher TD. 2009. Sexual conflict resolved by invasion of a novel sex determiner in Lake Malawi cichlid fishes. Science 326:998-1001. doi:10.1126/science.1174705

Science this week features an article by Elizabeth Pennisi about the research of evolutionary biologist Hopi Hoekstra. She studies pigment variations in wild mice.

Pigment clines have become an interesting model field study, because we now understand the molecular pathway of melanin production. With a dozen or so genes influencing natural pigment variations in mammals, it's a complex enough system that selection on color will lead to different genetic outcomes. That means we can look at parallelism in many natural cases to understand the evolutionary dynamics.

Hoekstra and her team are part of a genomics explosion in natural history studies. "This is an example of work ... merging the ‘green’ and ‘white’ side of biology, in which we learn about trait evolution from the biochemical levels within cells to how those traits are selected for or against in natural populations," says Hans Ellegren, an evolutionary biologist at Uppsala University in Sweden. Mark McKone, a biologist at Carleton College in Northfield, Minnesota, agrees: The work "could be a model for how to approach evolution in the postgenomic period," when genetic information and tools are more readily available.

A couple of weeks ago, Hoekstra's lab had a research paper illuminating pigment evolution among the deer mice of the Nebraska Sand Hills: "On the origin and spread of an adaptive allele in deer mice." I like this example a lot, because the color variation in Sand Hills is clearly postglacial -- this was not an agreeable Like the evolution of stickleback varieties in British Columbia, it's a good example of rapid selection on new colonists.

Multiple lines of evidence suggest that the wideband allele arose de novo and was not a preexisting allele. First, the haplotype carrying the deletion has greatly reduced variation relative to the wild type (Fig. 4A), which is inconsistent with a model in which the causative mutation was neutrally segregating in the population before any selective pressure (38). Second, the U-shaped SFS [site frequency spectrum] is most consistent with a model in which selection acts on a newly arising mutation (fig. S3). If the beneficial mutation existed on multiple haplotypes before the selective pressure, we would expect to see an excess of intermediate frequency mutations, which was not observed (38). Finally, the posterior probability density of the allele age falls entirely within the estimated age of the Sand Hills (Fig. 4B).

Taken together, our results demonstrate that variation at the Agouti locus is responsible for adaptive coloration in deer mice living on the Nebraska Sand Hills.

There are still details to be worked out in this example -- at the biochemical level, how does this allele cause lighter color? Can we say more about the spatial dynamics of the allele after it originated? But what I really like is that it shows obvious parallels to adaptive variations in humans that have recently been selected. It stands out in mice because Hoekstra is out there looking for pigment variations. Postglacial environments are one example in nature where recent environmental changes have generated new selection pressures. Of course, humans have induced new pressures on ourselves by means of massive cultural change.

Anyway, I thought it was worth pointing out the articles could go together as a set. I'll follow up with a second post on Hoekstra's take on developmental biology.

References:

Linnen CR, Kingsley EP, Jensen JD, Hoekstra HE. 2009. On the origin and spread of an adaptive allele in deer mice. Science 325:1095-1098. doi:10.1126/science.1175826

Pennisi E. 2009. How beach life favors blond mice. Science 325:1330-1333. doi:10.1126/science.325_1330

I feel like I've been transported into the future to see what science will be like fifteen years from now:

We successfully typed eight mutations in six genes (6) responsible for coat color variation for 89 [out of 152 tested; tables S1 to S5 (6)] ancient samples. To assess coat color variation of predomestic horses, we analyzed the bones of wild horses from the Late Pleistocene and Early Holocene found in Siberia, East and Central Europe, and the Iberian Peninsula. We found no variation in the Siberian and European Pleistocene horses, suggesting that these horses were bay or bay-dun in color.

...

In contrast, a rapid and substantial increase in the number of coat colorations is found in both Siberia and East Europe beginning in the fifth millennium B.P. (Fig. 1 and figs. S1 and S2). Although the earliest chestnut allele (MC1R gene) was identified in a Romanian sample from the late seventh millennium B.P., chestnut horses were first observed in Siberia (fifth millenium B.P.). Their prevalence increased rapidly, reaching 28% during the Bronze Age.

The whole paper is only a page long. Like, "Oh yeah, we just genotyped 89 horse skeletons from the Neolithic up to the Bronze Age, and here's how the process of selection on color patterns worked out."

Couple of things --

1. The pigment-altering mutations at these genes do not all show statistical signs of selection in contemporary samples of horses. But they aren't there in the ancient horses. That's the best evidence of selection you could possibly have. Message: tests of selection on contemporary samples are weak, particularly for loci with rare alleles or more than two alleles.

2. The study shows how much you can learn with a moderate number of samples. The advantages working in this case: the genes responsible for pigmentation phenotypes are well-characterized, the number of loci is well-matched by the sample size; the study focuses on recovering SNPs instead of sequencing.

UPDATE (2009-04-25): I looked at the statistical test of selection used in the paper with a little more detail. It comes from a paper by Jonathan Bollback, Thomas York and Rasmus Nielsen, published last year in Genetics. It's a very clever test. Assume a sample of genes taken from a population at some discrete set of times, t₁, t₂, t₃, and so on. Under genetic drift, this series of frequencies approximates a random walk, with the size of deviations depending on the effective population size. Under constant directional selection, the random walk will be biased in a way that can be approximated by a diffusion model of selection.

So if you have a big enough sample, you can estimate the relative contribution of stochastic (drift) effects and deterministic (directional selection) effects on the frequencies over time.

In the current case, two alleles have sample sizes that are large enough, frequency changes that are large enough and constant enough in direction to show that drift is minor and selection is strong. For example, the chestnut variant goes from zero in the Neolithic to 65 percent in the Medieval time period, with every change a between time increments an increase. That's an allele with 44 copies (by my quick count) in all samples.

Four alleles do not have such large changes (they are initially absent, but present in only four or fewer individuals in the later time intervals). There's only one copy of the SILV9 color variant mutation in the entire sample of archaeological horses. With a very small sample, the sampling variance will be larger than the stochastic effects of drift. So even if those few copies are exclusively in the latest time increments, the time series won't look unusual compared to drift.

But what the test doesn't consider is the current frequency. Since this is known with much less sampling variance, we can compare the present frequency with the frequency summed across the older time intervals to get a more powerful test of neutrality. Also, the test assumes that selection is constant -- if selection had a stochastic element, varying over time, that would have the same impact on the sample as drift or sampling variance. That's why the absence of an allele in an ancient sample can be stronger evidence of selection than the fine-scaled record of change over time.

Complicating matters is population structure. The coat color variants are not distributed evenly across the modern horse population; they are distributed into different breeds. The strong association of some color variants with breeds is, of course, evidence of its own that the color variants have been correlated with fitness under domestication. Now, whether this fitness association is a deliberate result of people liking colors (because they differentiate breeds, or look pretty, or whatever) or whether they arise incidentally (by linkage with other traits) isn't tested by these data. These functional aspects of selection provide another possible test of neutrality -- for example, in this case all the non-bay-black alleles increased over time, a bias that isn't consistent with chance.

References:

Bollback JP, York TL, Nielsen R. 2008. Estimation of 2N_es from temporal allele frequency data. Genetics 179:497-502. doi:10.1534/genetics.107.085019

Ludwig A, Pruvost M, Reissmann M, Benecke N, Brockmann GA, Castaños P, Cieslak M, Lippold S, Llorente L, Malaspinas A-S, Slatkin M, Hofreiter M. 2009. Coat color variation at the beginning of horse domestication. Science 324:485. doi:10.1126/science.1172750

According to this press release, gray hair in aging people is the result of a hydrogen peroxide metabolism gone haywire:

"Not only blondes change their hair color with hydrogen peroxide," said Gerald Weissmann, MD, Editor-in-Chief of The FASEB Journal. "All of our hair cells make a tiny bit of hydrogen peroxide, but as we get older, this little bit becomes a lot. We bleach our hair pigment from within, and our hair turns gray and then white. This research, however, is an important first step to get at the root of the problem, so to speak."

The researchers made this discovery by examining cell cultures of human hair follicles. They found that the build up of hydrogen peroxide was caused by a reduction of an enzyme that breaks up hydrogen peroxide into water and oxygen (catalase). They also discovered that hair follicles could not repair the damage caused by the hydrogen peroxide because of low levels of enzymes that normally serve this function (MSR A and B). Further complicating matters, the high levels of hydrogen peroxide and low levels of MSR A and B, disrupt the formation of an enzyme (tyrosinase) that leads to the production of melanin in hair follicles. Melanin is the pigment responsible for hair color, skin color, and eye color. The researchers speculate that a similar breakdown in the skin could be the root cause of vitiligo.

"As any blue-haired lady will attest, sometimes hair dyes don't quite work as anticipated," Weissmann added. "This study is a prime example of how basic research in biology can benefit us in ways never imagined."

Clearly this guy doesn't actually know any blue-haired ladies, most of whom are aiming for blue, to get rid of the yellowish color that may remain in gray hair.

But otherwise, I think this is quite cool information to have on this important variant of the pigmentation pathway. Age related-decline in two enzymes. There must be a bit more complication here than that -- for example, there is a huge variation among the population of follicles in the time of graying, even on a single person. There's also variation among hair types -- temple versus crown being an obvious example, but also beard versus crown. Not to mention others that I'd rather not mention...

The research does not investigate normal variation, just the metabolic mechanism. I'm curious about other primates. Silverback gorillas are the obvious analogy, but grayish or white pigmentation are by no means uncommon, and this mechanism provides another way besides mere pigment loss to get an age-related reduction in pigmentation.

(via FuturePundit)

Mark Derr of the NY Times reports on a new study showing that black North American wolves got their melanism from dogs:

In a bit of genetic sleuthing, a team of researchers has determined that black wolves and coyotes in North America got their distinctive color from dogs that carried a gene mutation to the New World.

The finding presents a rare instance in which a genetic mutation from a domesticated animal has benefited wild animals by enriching their “genetic legacy,” the scientists write in Thursday’s Science Express, the online edition of the journal Science. Since black wolves are more common in forested areas than on the tundra, the researchers concluded that melanism — the pigmentation that came from the mutation — must give those animals an adaptive advantage.

There are so many examples of this phenomenon in mammals now! This one is interesting because it would have been carried in by early dogs brought in via Beringia -- so it's another case where an intercontinental migration has brought a new adaptive allele that introgressed into a natural population.

There is also a date:

Comparing large sections of wolf, dog and coyote genomes, Dr. Barsh and his colleagues concluded that the mutation arose in dogs 12,779 to 121,182 years ago, with a preferred date of 46,886 years ago. Since the first domesticated dogs are estimated to date back just 15,000 to 40,000 years ago in East Asia, the researchers said that they could not determine with certainty whether the mutation arose first in wolves that predate that time, or in dogs at an early date in their domestication.

This could have been selected in the very earliest domesticated dogs, based on that date. It would be useful to have a number of genomes from ancient wolves to screen against variation present in the wild population around the time of domestication.

The really cool thing is that we will probably have samples like that within the next several years...