Archive | Eye color

Some new research into blue eyes and KIT mutations

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A tobiano pony with partially blue and partially gold eyes (cause unknown)

A year ago I posted expressing my doubts about the theory that blue-eyed horses with KIT mutations (tobiano, sabino and the white spotting patterns) must carry an additional mutation to account for their eye color. At the time, I did not think that there was enough evidence to merit the absolute terms that were often used regarding this theory. It is also true that, having tracked so many instances of blue eyes in the patterns in question, I thought the weight of probability favored the theory that the blue eyes were part of at least some of these patterns and not always a separate mutation. At the time I wrote:

The common theory in horses is that these W-series horses must have a splash mutation as well. And they may. Certainly there are far more mutations for white patterning than previous expected. I have long thought that the numbers of blue eyes on the dominant whites, particularly among the founder horses (ie., the horse that carried the initial mutation) were just too high for them all to happen to have a splash mutation as well. I did not have an exact number, though – just a sense that it was high. But the new sort gave me a number – six of twenty.

The argument that KIT mutations are “incapable” of producing blue eyes is based on information about how pigment is formed in the eyes of mice. I found the theory difficult to evaluate because the passages referenced by its supporters do not deal directly with eye color in horses, and I simply do not have the deep level of understanding of eye structure necessary to extrapolate beyond the specific subject, which was not the absence of blue eyes but the presence of dark eyes in a particular mouse color.

But perhaps more importantly, I just wasn’t sure that mice and horses were the same in this regard. In fact, I just wasn’t sure that eye color in various mammals might not be different in significant ways, just as other aspects of coloration vary between species. In that same post, I included a picture of what was then a newly-identified KIT mutation in dogs – the panda pattern in German Shepherds. I found the blue eyes on the founding dog particularly compelling because in dogs blue eyes are not generally associated with the common forms of white patterning, despite the fact that the mutations for most of those patterns have been found on MITF, which those of us more familiar with horses think of as the “splashed white” gene.

It seemed to me that these blue-eyed (KIT) German Shepherds, and the more common dark-eyed “irish marked” (MITF) dogs were a pretty good argument that when it came to eye color, there was probably some variation from the mouse model.

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The same pony, with one predominantly blue and one
predominantly gold eye (and an occluding spot over his blaze)

Some time after that post was made, I ran across a paper on Blue-Eyed White (BEW) alpacas. In that paper, the blue-eyed white phenotype was linked to the presence of two mutations to KIT. The alpacas, which were uniformly blue-eyed, were compound heterozygotes for two different white patterns (bew1 and bew2), both located at KIT. Since that time I have been able to confirm with one of the researchers that there were no mutations at the sites associated with splashed white in horses that could be correlated with the blue eyes on these alpacas.

Then just this month a paper was published linking the blue-eyed white phenotype in cats – called Dominant White (W) – to a mutation on KIT. In that study, the authors were quite clear about the connection between KIT and blue eyes.

In the population sample, we were also able to examine the correlation between genotype at the W locus and iris color. An individual that is homozygous W is much more likely to have blue iris, exhibiting odds 77.25 times larger than the odds of having blue irises of a genotype other than W/W (p < 0.0001).  An individual that is heterozygous (W/w+) also demonstrates increased odds of having blue iris (OR=4.667), four times larger than the odds of having blue irises of a genotype other than W/w+ (p=0.046) The odds of having blue irises of a wild type individual is 0.

With those two studies seeming to cast considerable doubt on the “Never From KIT” theory, I decided to contact one of the corresponding authors with some questions in hopes of getting a better understanding of this topic and of eye color in general. What I was told was that eye color is most likely a polygenic trait, and that it really does depend on the species, as well as the specifics of each particular mutation to KIT. That could explain why some KIT mutations are more prone blue eyes than others, as well as why there appears to be a higher incidence of blue eyes in homozygotes of some patterns.

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On this eye, the blue and gold portions are interspersed in such a way that the colors appear softly blended

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On this eye, there are fewer flecks of blue, as well as irregular patches of dark brown

So why does it matter if KIT mutations can produce blue eyes alone, or if they need a splashed white mutation? What purpose does this kind of information serve? The fact is that knowing the cause can help breeders more reliably get the outcome they desire, whether they wish to breed for or select against blue eyes. Likewise, breeders seeking to produce – or avoid – the splashed white phenotype need to know if blue eyes are always significant. If it is possible for some of the other patterns to produce blue eyes independent of a splashed white patterns, then assembling a herd of blue-eyed tobianos in hopes of developing a line of splashed whites is going to ultimately prove frustrating.

It is also true that quite a few blue-eyed horses have come back negative for the known splashed white patterns. It is likely that some of them have as-yet-unidentified splashed patterns. However, if some have blue eyes that are just a less common aspect of a pattern that is already identified, then knowing this could spare their owners time and money spent looking for something that is not really there. So while the subject is quite technical, it really does have a very practical aspect.

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Range of expression for classic splash

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One of the big questions about patterns is the outer limits of their expression. That is, how much or how little white can there be in any given pattern? Some patterns, like those in the W-series (“dominant white”), seem to show a lot of variation both in the extent of the white and the overall look of the pattern. Other patterns have a little more consistency in their expression. Consistency is what makes a pattern like tobiano so much easier for the casual observer to identify. But even relatively consistent patterns do have a range, and individuals that deviate quite dramatically from what might be considered typical. Animals at the extreme ends are particularly interesting because they tell us where the white most likely originates for that pattern, as well as how smaller-effect modifiers (and possibly other patterns) interact with it.

Classic splash – that is, SW1 in its homozygous form – is one of the patterns that is prone to a fairly consistent expression. That was why it was fairly easy to identify the presence of that specific pattern in breeds prior to the advent of genetic tests. Homozygous SW1 horses had a consistent look, and that look was pretty distinctive. Even so, there were horses that fell along the more minimal end of expression, particularly among some of the rustic pony breeds where white markings tended to be minimal or absent altogether. These horses skirted the edge of the loose rules I had for identifying the classic splash pattern: four white legs, some white on the lower body, white on the tail tip, white face, two blue eyes. Some tended to blur the line between what I thought of as a sabino-boosted heterozygous SW1 and other splash-like patterns (what turned out in many cases to be SW2). Testing has now made it easier to determine when horses from that latter group really are carrying SW1, but I did wonder what testing would tell us about how far thea “rules” could be bent – or if they could be broken altogether. Tobiano testing confirmed that pattern’s biggest rule – four white legs – could be broken. I wondered if what I thought of as the biggest rule for homozygous SW1 – two blue eyes – was also negotiable.

So I was thrilled when Pauline Högdahl sent a photo of Stenbydal’s MJ, the Gotland Pony colt pictured in this post. His owner, Emmelie Nilsson, had him tested and he is in fact homozygous for SW1. To date there have been a number of whole-colored (ie., no white markings whatsoever) heterozygous SW1 horses, but this fellow is the most minimal homozygous SW1 that I have encountered.

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He does have white on all four legs, though the marking on the right front is pretty minimal. His tail end is white, and he does have a small patch of white on his belly. 


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But remarkably enough, he does not have blue eyes. He does have a small blue segment in one eye, but the eyes are otherwise dark. So while very white faces and blue eyes are often considered the defining characteristic of a splash white horse, and homozygous SW1 horses display the classic form of the splash white pattern, having two blue eyes is not an absolute, unbreakable rule. 

While Stenbydal’s MJ may answer the question of whether or not homozygous SW1 horses have to have two blue eyes, he poses new questions. What exactly causes a pattern to be so minimal? Is it all random chance? Or is there some genetic component that tamps down the white, just as there are genetic factors that amp it up on some horses? The latter does seem more likely, especially given that minimized patterns tend to run in families. Here is a picture of MJ with his dam, who would presumably be heterozygous for SW1. 

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His sire, Skotte 492, would also carry SW1, but only has a few white hairs on his forehead. He has at least one other tested homozygous SW1 that is quite minimal. That is Ängdalas Spirello. He also has a son, Zidane Zojvide, that has what appears to be a classic pattern but with one dark (or at least primarily dark) eye, but I am not aware of any test results on him. (More photos of Zidane Zojvide as a young horse, where the contrast on his pale palomino coat is a little better, can be seen here.)

This question of factors that suppress patterns, or boost them, ties into some of the new research on white markings. That is a subject I’d like to cover, especially in light of the new paper that was published just recently. To do that, however, I want to give some background on past studies of white markings, including what was already known, and the system that was developed to begin assessing white. Watch for those posts in the upcoming weeks, but I have at least one more splash white post to make first.

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Musings on blue eyes (again)

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The blog here continues to be somewhat silent while I work on the next book, “Equine Tapestry – An Introduction to Colors and Patterns“. What began as a reissue of the front portion of Volume I, Draft and Coaching Breeds in color has (predictably) taken on a life of its own. The original text touched on the as-yet-unidentified pinto patterns, with an attempt to classify the different categories of sabino-like patterns visually. Since that time, quite a few more pieces of the puzzle have fallen into place, and that has lead me to do a major overhaul on that material.

One of the things that became clear to me with this new information was that the way I personally organize my files was leading to blind spots in my understanding. I have mentioned before that a huge part of my research involves massive notebooks with images and pedigrees. Because I am a visual learner, sorting things this way helps me make connections that I might otherwise miss.

Notebooks

I have organized information in notebook pages like these for more than 20 years. Soon after I began, I started sorting the information by breed. Because so much of my interest centered around which colors were present in which breeding population, this made sense. What I began to suspect, working on the new book, was that I needed to rearrange some of my notes by color groups, rather than by breed. The previous structure was great for seeing how some of the louder sabino patterning arose in the Arabian breed. Seeing how that worked convinced me that the louder horses were some kind of new mutation, and not just a more extensive expression of the existing “flashy white” in some lines. Laid out in a breed-centered notebook, it was clear how these louder horses – horses like Rhocky Rhoad – might come from flashy families (like Khemosabi), but their (numerous) relatives did not generally look unusual, while their own descendants most certainly did.

But this same structure made it harder to make connections about the colors themselves. The sheer volume of information – I have thousands of horses on file, and many times that number waiting to be included – made those connections more dependent on my working memory. Writing more has meant less time for musing, and with it the chance that I would make those mental jumps. When recent papers made it clear that my hunch about horses like Rhocky Rhoad were correct, I decide it was time to set up a parallel set of notebooks for the pinto pattern categories, starting with the W-series (W1-W20). I was hoping this would give me better insight into how the patterns within that group – the group previously called “dominant white” – worked. With luck, I might stumble upon the best way to present this very varied, and not-entirely-helpfully-nameed group in the new book.

I now have all twenty of the known W mutations, along with images of every known or suspected carrier, in a single notebook. The first thing that jumped out at me was something that has been a bit of a hobby horse for me for some time now. That was how many of these lines involved blue eyes. Of the twenty families, six have blue-eyed individuals. In some cases, like the W5 family member Sato (above), just have a blue segment. Others have a full blue eye, or even two blue eyes, thought that last is actually pretty rare.

This is not surprising to anyone who has looked at historical records of white-born horses. Blue eyes are not infrequently mentioned. They are mentioned in connection with some of the old European studs that previously bred white-born horses. Early researchers also comment on their occasional presence. Nowadays, a search on the internet will turn up any number of commenters that will tell you that this “obviously” means the horses carry a splash pattern. You can even find those who will assert that no KIT mutation ever produces blue eyes in a mammal.

That was part of why I included the image of the panda German Shepherd a while back. She has a newly-identified KIT mutation, and she most certainly has blue eyes.

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Interestingly enough, most white patterning in dogs has proven to be caused by mutations to MITF – not KIT. In horses, MITF is the gene associated with splash patterning – and with blue eyes. For those familiar with dog coloring, the “extreme piebald” found in many sporting breeds, the “color-headed white” pattern in Collies and Shelties, and many forms of “Irish Spotting” have all been mapped to MITF. None of these patterns is associated with blue eyes in dogs. There is a MITF mutation in dogs – the one that produces white in Boxers – that produces blue eyes on rare occasions, to this is not an absolute, but generally speaking these MITF mutations are not associated with blue eyes. (For more information on the different MITF mutations in dogs, this a good site.)

The common theory in horses is that these W-series horses must have a splash mutation as well. And they may. Certainly there are far more mutations for white patterning than previous expected. I have long thought that the numbers of blue eyes on the dominant whites, particularly among the founder horses (ie., the horse that carried the initial mutation) were just too high for them all to happen to have a splash mutation as well. I did not have an exact number, though – just a sense that it was high. But the new sort gave me a number – six of twenty. That’s a lot, especially with breeds where there is no evidence of the presence of the one splash pattern (classic splash, SW1) known to remain cryptic in its heterozygous state. The other “new” splashes are dominant mutations, and a good bit more obvious in terms of phenotype.

What is interesting is that the one horse that is often asserted by online commenters to “surely be a splash” is the well-known Arabian stallion Khemosabi. This is based on the fact that he has multiple blue-eyed descendants. What became clear as I resorted these records was that yes, he does have a number of blue-eyed descendants. However, all those in my files are also members of the two W-series mutations that occurred in his line. One would expect, if Khemosabi carried a splash mutation (at least, as we currently understand the pattern), it would appear in more than two lines. He did, after all, sire over 1,250 foals. Finding it in two lines, which also just happen to be those that have formally identified white spotting mutations, seems to suggest that the blue eyes are part of the pattern and not some additional inherited trait.

There are caveats to this, of course. Blue eyes are notoriously underreported. When I began to suspect that the louder sabino expressions in Arabians were new mutations, I printed out the markings files for the families where they occurred. So in my files are the marking diagrams for all the first generation Khemosabi descendants. It is quite possible that there are horses in that group with blue eyes that were not part of the registration records. Blue eyes in Arabians are still considered a serious fault, so there is some incentive to overlook it when filing a description, especially for an eye that is not completely blue. (I am always looking for images and records of blue eyes in Arabians, if anyone has them, by the way!)  I cannot also be sure that some of the other lines – the fourteen that are not included as having blue eyes – might not also have blue. In many cases, eye color is not mentioned at all,  and there I have defaulted to the assumption that the eyes were dark. That is not the same as knowing the eyes are dark, though.

My next task is to assemble the even larger group of suspected dominant white horses into one notebook. When that task is complete, I should be able to do another post about the status of the blue eyes in that group as well.  Well, that and get a little closer to a finished book!

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