Q?rius Presentation: The Dog Genome: Shedding Light on Human Disease – Elaine Ostrander

[applause] Elaine Ostrander:
Hi, everybody. So thank you so much for coming. So for the row of people sitting behind me,
you do not get to fill this out until you’ve heard me speak. So put your pens and pencils
down there, all right? So how many — how many people in the audience have dogs? Yeah,
so good this is a great, dog-friendly audience, because we’re going to be talking today about
dogs, and a couple of different aspects: we’re going to talk about health as well as morphology,
you know, why does are different shapes, and how all that data sort of comes together to
inform us about human health and human biology as well. So we’ll leave some time at the end;
we’ll have lots of time for questions. So this is the closest ancestor to the dog. Who
knows what we’re looking at here? Audience Response:
Wolf. Elaine Ostrander:
What kind of wolf? Gray wolf, right, of course. So dogs were believed to have domesticated
from gray wolves. The latest data suggests maybe around 13,000 years ago. So evolutionarily,
that’s really a — that’s a drop in the bucket. I mean, that’s not very long to go at all,
and so one of the things we’re always sort of thinking about and pondering about when
we look at our pets, wandering around our house is, you know, is whatever they’re doing
caused by genes that are embedded in this wolf genome, or is this something new that’s
come up during the process of domestication? And that’s one of the things that we go back
to and we ask ourselves over and over and over as we look at all of our friends. So how many of you recognize your dog breed
up there? All right, there’s got to be some golden retriever owners, I’m betting? There’s
probably some schnauzer owners, right? Maybe some hound owners? You know, a Weimaraner
owner or two? So these are a small number of the 175 breeds of dog that are recognized
by the American Kennel Club. The American Kennel Club is the largest registering body
of dogs in the United States today, but they are not by any means the only registering
body. There is the United Kennel Club, and several other kennel clubs as well, and there
are kennel clubs actually all over the world that in aggregate recognize 493 different
dog breeds today — 493 different dog breeds today. And those dog breeds show an extraordinary
amount of variation, right? So we would say that there’s probably three things you need
to keep in mind as we’re going to talk about dogs for the next 45 minutes, or so. It’s
important to remember that, even though all of these guys look very, very different — they
have a different body size, different coat colors, different head shapes, different leg
lengths — all dog breeds are members of the same species. So they all have the same karyotype
or the same chromosomal organization; they all have the same genome organization; and
they can be crossed to do produce fertile offspring. Now, clubs like the American Kennel
Club don’t really encourage you to cross dogs from one breed to the next, and as a matter
of fact, to be, for instance, a registered golden retriever, your parents had to be AKC-registered
golden retrievers, and your grandparents in turn had to be AKC-registered golden retrievers. So one of the reasons people like me study
dog breeds is because each one represents sort of a closed population. So if you think
about human populations that are isolated, you know, people who live in Finland or Iceland,
or people who have lived on islands for many, many generations, or the Bedouin pedigrees,
geneticists like to study those kinds of populations because there isn’t a lot of add mixture from
the outside. They really have sort of a set number of different alleles at each genetic
loci, and that makes the problem of studying complex traits, like diabetes, and cancer,
and epilepsy actually an awfully lot easier. The only problem in human genetics is that
there’s a limited number of such isolated populations, whereas in the dog world we have
493 of them. And so we actually in my lab have been working hard to get DNA samples
from each one of those 493 breeds. We have a great relationship with the American Kennel
Club. We don’t breed any dogs. We don’t keep any dogs in kennels, but if you go to a dog
show or an obedience trial, or a specialty or an agility trial, anywhere here in the
tri-state area, you’re probably going to find someone from my lab there taking cheek swabs
or handing out kits or collecting blood samples. And we’ve now got a set of 50,000 DNA samples
in my laboratory. So that’s pretty impressive, huh? So when we look at these dog breeds, we really
see extremes of variation, and one of the things that makes a breed a breed is that
that variation breeds true. So if you cross one golden retriever to another golden retriever,
the puppies are all going to pretty much look the same, and there’s some nice examples up
here for you. You know, if you cross one Dalmatian to another, they’re all pretty much going
to look the same. Cross one boxer to another, they’re all pretty much going to look the
same. And the American Kennel Club is really strict about the things that are important,
and that define each breed. And it’s different for different breeds. For some breeds it’s
their coat color; for some, it’s how tall they are; for some, it’s how long their legs
are; for some, it’s the shape of their face; how far apart their eyes are; whether their
ears are perked or whether they’re down; and when a dog goes into a dog show, those are
all the things that the judges are actually grading them on. So because breeders have
been breeding for those traits for years and years and years and years and years, those
are the things that in my lab we’re trying to find the genes that control them, and in
doing so, we’re sort of getting a vocabulary of growth regulation that we really can’t
get from studying worms, and flies, and mice, and rats, and all those other traditional
model organism systems. Now, I really like this picture, and don’t
worry if you can’t read the writing here, but what we’ve done is we took 1,000 DNA samples
from dogs representing 85 different breeds. So we took about 12 different dogs from each
breed, and we tested their genome at 100,000 different positions, and we were looking at
the variation in the A’s and those T’s and C’s and G’s that you’re always hearing about.
And then we fed it all into a computer program, and we said, tell us how all the dog breeds
relate one to another; and that’s what this wheel — and you can really follow the color
coding in the round — is telling you so well. So if you look at your upper right, you see
there are the red, and you see spaniels, the American cocker spaniel, the English cocker
spaniel, the English springer spaniel, the Cavalier King Charles spaniel, the Irish water
spaniel, the Brittany spaniel, and so on. And if you look over down at about 4:00, you’ll
see the sort of bull mastiff-type dogs — the miniature bull terrier, the French bulldog,
the bulldog, the boxer, and so on. Now, we have this hanging in my lab and we
use this every single day as we’re developing a hypothesis. So if I’m going to study something
like epilepsy, and I’ve got a whole bunch of samples from English springer spaniels
— those are the red up there at about 2:00 — then I can actually probably go out and
get DNA from affected American cocker spaniels, and Irish water spaniels, and Cavalier King
Charles spaniels that are also affected, and I can probably be correct when I hypothesize
that they all got the disease because they carry a mutation in the same gene. And that’s
because they all share a common set of founders; that’s why they’re all colored in red. And
I study epilepsy in the mastiff-like breeds — well, they probably have a mutation in
a different gene, but when I look at the mastiff, and the bull mastiff, or the bulldog, or the
boxer, and the French bulldog, again, they’re all colored in kind of that teal color. They
probably got the disease because, again, they share a mutation in the same gene, and they
likely got it from a founder. So how long ago did this founder live? Well,
how long ago do you think most dog breeds were developed? You know, domestication occurred
about 13,000 years ago. How long do you think most dog breeds have been in existence? Any
guesses? Female Speaker:
Two hundred. Elaine Ostrander:
You’re pretty close. About 300 years. So most dog breeds were developed in Europe, most
were developed by fanciers during the Victorian times. So most of them have only been around
really for 200 or 300 hundred years. So again, evolutionarily, we’re not even talking about
a drop in the bucket; we’re talking about a, you know, a perspiration drop in the bucket.
I mean, as tiny as you can really get. So we can make these kinds of hypothesis when
it comes to a morphology or when it comes to disease susceptibility, and we’re usually
right. And the other thing is that the genes that we end up finding by studying dogs usually
turn out to be important for the same things in humans. It’s just that, gosh, it’s a lot
easier to go to my freezer of 50,000 dog DNA samples, and pick out a few related breeds,
and look up their health records, and find out what they’ve got and what they don’t have,
and correspond with their owners, and start studying a pool of affected and a pool of
unaffected dogs in order to find a disease gene of interest. Okay, so this is actually one of my favorite,
favorite dog pictures. This is a Harlequin Great Dane. It’s skeletally among the largest
of the dog breeds, and down here is a little Chihuahua, which is skeletally among the smallest
of the dog breeds. Now, we’ve been studying dog morphology for several years in my lab,
and we’ve published papers describing genes that control body size, and leg length, and
skull shape, and fur, and we rely on samples from these extremes in the population in order
to do studies on things like body size. Again, remember, both of these guys are members of
the same species, and they could be crossed — these two guys here — to produce fertile
if singularly unattractive, but nevertheless fertile offspring, and you might want to give
a little thought as to who would be the male and who would be the female when you, you
know. All right, so what do we have over here? [laughter] Elaine Ostrander:
Okay, so this is, again, a Chihuahua; this is Zeus. I had to put this picture in. Zeus
actually holds the 2013 Guinness Book of World Record as the tallest dog. So he is 44 inches
at the withers, or the shoulders. This is not photo shopped. This is actually how tall
Zeus really is. And so we’ve been collecting samples from dogs at these extremes, as well
as everything in the middle, in order to fine genes that are responsible for this dramatic,
dramatic difference in body size that exists within this one species. And I can tell you
from a couple of papers that we’ve published that there are about a half a dozen genes
that account for most of that size variation. And so I didn’t put their names up, but if
you’re interested I can tell you they’re the insulin-like growth factor I gene; there are
a couple genes on the X chromosome; there’s the insulin growth factor receptor; [unintelligible],
HMGA 2, STT 2, growth hormone receptor. And so some of these may sound familiar to you,
because they’ve been shown in studies of mice to be important in controlling mouse body
size. The only thing is you don’t learn a lot from studying mice in this case, because
you don’t find mice that differ in size by 40 fold, right? I mean, wouldn’t that be truly
frightening, I mean, if you did, right? But of course, we do find that in dogs. So we’ve spent a lot of time identifying the
genes and the mutations that account for variation in body size. And we just published this,
and I know it’s kind of complicated, but I wanted to show it to you, because I’m actually
really excited about the implications of this study. So if you look on the left, you see
a bunch of bricks that are either yellow or red, and I know it’s probably hard to read
on the bottom, but on the bottom is the name of each one of these genes; one at a time:
growth hormone, IGF 1 — so on until we get to IGF 1. And you don’t have to worry about,
you know, really which gene is which. Going along the vertical are the weight in pounds
of different dog breeds that we’ve assayed. And then what the red and the yellow is telling
you is that at each one of these genes, there’s two possible alleles, or there’s two possible
mutations or variants that can occur. One is an ancient variant that we find in wolves,
and one is a new variant that’s only been on the planet for a few hundred years, and
we only ever see it in domestic dogs. Now, the yellow is the ancient variant that we
find in wolves, and red is the new variant that we’ve only ever seen in dogs. Now, what’s
really striking is if we look at really little dogs that weigh like zero to five pounds,
all the way across you see red, telling us that those dogs have the new variant, the
one that we only see in dogs at every single one of these genes, but as dogs get bigger
and bigger and bigger, that drops off until it’s basically mustard yellow. And so all
those big dogs are big because they carry the DNA variant that came from the ancient
wolf. So that’s kind of cool. So we took that data and we said, how predictive
is it? You know, if I actually take what the data’s predicting versus what I really see
in a panel of 500 dogs, how good is the predictive value of my half a dozen genes? And that’s
what this line over here on you right is telling you. And the fact that you get a pretty good
line from all of those data points is telling you just those six genes, if we could assay
them in every single puppy that was born, would have actually have great predicative
value for what the final size of the dog is going to be. We’d be right to about 82 percent.
So think about this. This means that you could go to the pound, you could get a puppy, you
could get a cheek swab, you could get it assayed at these six genes, and you would know what
the final size of that puppy’s going to be pretty close to accurate, to about 82 percent
accurate. So that’s really amazing. Six genes, just six genes control that much variation. Now, one of the things about this study is
it actually only holds true for dogs that weigh up to 90 pounds, and if you think about
the giant breeds — the Great Danes, the Newfoundlands, the Saint Bernards — these six genes only
have about 5 percent predictive value. So there’s probably lots of genes responsible
for giantism [phonetic sp] in dogs that I actually haven’t found yet, and that’s one
of the things that my lab is in the process of doing so that we can extend this line,
and make it bigger and bigger and bigger until we can actually understand all the genes controlling
that full range of body size, going all the way up to, you know, 180, 200 pounds. Now, as we’ve begun to publish more and more
of this data, people have gotten really excited, and they’ve kind of picked out different things
that they want to study, or that breeders would like to try and breed for. So I have
people in my lab studying leg length, and studying skull shape; others are continuing
to study body size, some are even studying performance. And we built a big dataset based
on 1,000 different dogs from 85 breeds, and we also included 500 wild canids — so coyotes,
wolves — from all over the world. And we tested their DNA at 1,000 different points
in the genome, and we made that DNA publicly available without any restrictions, without
any patents to anyone who wants it. So you know, we really encourage other people to
try and think about these problems as well. Now, I like this particular story. This is
one that was led by Heidi Parker [phonetic sp], and she was a graduate student in my
lab, and then she went on to be post-doc, and then she went on to be a staff scientist,
and she’s been with me for 15 years, and I actually don’t think she’s ever going to leave.
So Heidi has been really interested in short-legged dogs. And there’s about 20 such breeds. They’re
called chondrodysplastic. They have a ratio of height to body length less than zero. So
they sort of have a normal head, and a normally proportioned body, but then they have these
very short and thick legs. In the dog world, we would say their structure is well-boned
or heavy, and their forelimbs, like you can see on that basset hound, are often sort of
bowed, or a little bit curved out. So Heidi came to me one day, and she said
“I want to try and find the genes responsible for this trait across these 20 breeds.” And
I said, well, you know, this could be a really hard problem. It could be that they all share
a common mutation, but these breeds were developed for different purposes. Some were developed
to be — to go down rabbit holes; some are fox hunters; some are companions; some are
ratters. So I said I’m not so sure they’re all going to have the same mutation in the
same gene. And she said, “Well, I think this is a really interesting problem; I’m going
to try it anyway.” And she’s not being totally unbiased here, because you see that doxen
and that basset hound? Those are actually her dogs. [laughter] Elaine Ostrander:
Yeah. And so, you know, when you come to my lab and you work, people are often motivated
by things that they have with their own dogs, and you know, some of my graduate students
are top — have top show dogs, or top dogs in the agility ring. So it’s not — I’ve actually
had mushers come and be graduate students in my lab. So, again, motivated by trying
to understand the underlying genetics. So Heidi went after the gene for chondrodysplasia.
Now, I know this is — looks like just a bunch of lines to you, but what you’re looking at
here in the alternating gray and black, each one represents a different dog chromosome;
and dogs have 38 chromosomes, and we also included the X, but we didn’t put in the Y,
because we figured there was nothing important on the Y anyway. And then what we’re doing
is we’re comparing cases, who are chondrodysplastic, to what we call control, which are all the
other breeds that are not chondrodysplastic; and we’re looking at 100,000 different points
in the genome, and we’re saying, is there a chromosome that has a data point that’s
significantly different between cases and controls? And you can see this is very significantly
different here on canine chromosome 11. And for those of you who’ve calculate P-values,
the P-value here 10 to the negative 102. So .0000000 — put 102 O’s, and then a one, and
that’s how statistically significantly this is. So this is hugely statistically significant. So we decided to follow up on this, and the
way we did it is we looked for something that evolutionary biologists call a selective sweep.
So what’s a selective sweep? Well, when you have a selective sweep, you assume, as I’ve
told you before, that there’s an ancestral mutation that occurred many, many generations
ago. In this case, before dogs were divided up into lots of different breeds. And then
dog breeders breed, and they breed, and they select for different things, and there’s a
lot of scrambling of chromosomes, but the mutation stays, because they’re always still
selecting for that one trait; in our case, chondrodysplasia. And so now, when we look
at modern day dogs, their chromosome may look nothing like the ancient chromosome, except
for where the mutation is, and in the space right around the mutation. And so we look
for that region of commonality, and when we find a region of commonality across a group
of breeds who have a trait, then we know the mutation has to be somewhere in there, and
that turns out to be exactly the case here. So this is an old fashioned gel, and I put
up because I think probably some of you in college had a — have had a chance in science
classes to run a gel. A gel simply separates DNA based on its size. The control dogs are
things like greyhounds, and boxers, and cocker spaniels, and if you look at the top, you
don’t really see anything of the gel. But when you look at the case dogs, and these
of course basset hounds, and the doxens, and the Pekingese, you see a bright yellow — a
bright white band. And that’s telling you that all of these cases have some extra DNA
that’s responsible for this trait that’s not present in the group of controls. So right
away we know that what our mutation is — it’s not a single base pair change, and it’s not
a loss of DNA. All these chondrodysplastic breeds have acquired extra DNA. And in fact,
they’ve acquired an extra copy of a gene called fibroblast growth factor IV. Now, they didn’t acquire any of the regulatory
machinery that tells it when to turn on or when to turn off, but they acquired the full
sequence of the gene. And so actually the genes around it, they sort of parasitize the
regulatory machinery from genes around this — what we call retro gene, and that’s telling
it to be expressed in fetal chondrocytes, and you know what that’s doing? That’s closing
the growth plates prematurely. So the legs never elongate as long as they should. This
gene is expressed, the growth plate closes, and er, er, er, er, er [phonetic sp] — the
leg can’t elongate to its full and natural length. And every single one of those 20 breeds
I showed you has exactly that gene, including this, which is the corgi breed. So this was really exciting, and we were able
to publish this ‘Science,’ not because the editors of ‘Science’ care about corgis, although
I think they should, but because this was sort of a new way to screw up the genome that
had never really been described before in mammals. And the other thing is, we of course
know that there are humans who suffer from forms of what we’ve historically called dwarfism,
but it’s really chondrodysplasia, and we don’t always know what’s causing that. So now this
gene goes into that lexicon, into that vocabulary as something that we need to think about when
we look at those. So this is a really neat example, just by studying a phenotype that
dog breeders have been breeding for for a couple of hundred years. In a bunch of healthy
dogs using DNA from my freezer, we’ve been able to figure out a whole new mechanism for
screwing up the genome, and we’ve been able to add a gene to the medical genetics vocabulary
that turns out to then become very important. So this to us is really a huge success. So we’ve gone on to do that in several other
ways, and I’m going to give you one more example in morphology, and then I’ll give you one
example in disease, and then we’ll have some time for questions in the end. What are these? Audience Response:
[inaudible] Elaine Ostrander:
Absolutely. These are all skulls from dogs, and these are pictures we took down here at
the Smithsonian. Turns out they have a lot of skulls in the back room. [laughter] Elaine Ostrander:
Right. And these are all different dog breeds, and they differ in both shape, and what else? Audience Response:
Size. Elaine Ostrander:
And size, exactly. So when Jeff Shaunenbeck [phonetic sp] joined my lab, he said “I want
to find the genes that control this. I want to understand the genetics of the skull shape
and size in dogs.” And I said, Well, I don’t know. This sounds like a really hard problem,
but Jeff had one of these giant Leonberger dogs with a big round, kind of fluffy face,
and so, you know, this is what he wanted to do, and so that’s what we did. Now, the first
problem we had was we don’t know how — wow do you quantitate a skull, right? I mean chondrodysplastic
is easy. The dogs either have it or they don’t have it; and body size is easy — you measure
them or you weigh them. How do you actually quantitate a skull? It turned out to be a
hard problem, but we solved it, and what we do is we have something called a MicroScribe
Digitizer, and we touch each skull at 51 different landmarks, and that sends data to a computer,
and then in the end, the computer takes the 51 data pieces and it draws a three dimensional
picture of what that particular skull looks like. So here I’m showing you top and bottom and
side views of a particular dog’s skull, and everywhere there’s a red or a blue number,
that’s one of those data points that we’ve gotten. So the palate can be long or short
— that’s the roof of your mouth — for a dog that has a long or a short snout; that
angle in picture C between the rostrum and the nose, that can be, you know, pretty much
a ski slope, or it can be at a right angle, like it is in a Newfoundland. We would say
they have a Roman nose, or a very high forehead; and there’s variation actually in every one
of these traits. Now, we’ve actually been fortunate to travel around the world. We’ve
measured about 1,000 skulls from 161 different breeds, and people always ask me, where do
you go to do this? And we go to lots of museums, and certainly the Smithsonian is the first
place we went, but we’ve been to museums and universities all over the country, and actually
all over the world. The university in Switzerland actually has something like 2,000 canine skulls
that we’re about halfway through measuring now. But I have to admit, there are a group of
people in the United States that have a lot of skulls in their basement, and I don’t why,
but they do. And they all call me, and they say, you know, I got a bunch of dog skulls.
Why don’t you come down and measure them? This is Alex, one of the people in my lab,
and she’s measuring the dog skulls. This is a gentleman in California, who called us to
fly out and go down to his basement, and measure all the skulls, and you can see he has all
kinds of animal skulls. We really don’t know why. We don’t ask those questions. We just
measure the dog skulls, and get out of there, but yeah, there’s a lot of these people in
America. They all want to friend me on Facebook. You know, it’s a whole culture thing. So anyway,
but these people have been very generous with their collections, and we’ve actually gotten
a lot of data from skulls where we could verify the breed and we could verify the age. Okay, so this is — this is in some way sort
of a tragic slide. So this is really what motivated Jeff to begin this project. So in
the left-hand column are a set of human conditions that are really different kinds of craniofacial
abnormalities, and I know not — you’re not familiar with most of those words, but they
describe different abnormalities that we see unfortunately in humans, often associated
with particular syndromes. And next to them are breeds where the breeders are actually
trying to breed that in as part of the breed standard, and really one of the examples that
I use — and this is not something that the American Kennel Club is doing, this is not
an AKC breed, but this dog shown here is a Pachón Navarro, and the breed standard includes
having this deep cleft that goes all the way from the outside of the snout, all the way
down in to the roof of the mouth. So you know, this is not something that American breeders
are advocating, but you know, it is something that, you know, we see and so we want to try
and find those genes, because we think that’ll help us understand something about cleft lip
and cleft palate, and we’re actually interested in all of these craniofacial features. So the one that we’ve been doing the most
work with is called brachycephaly, and that means having a very pushed-in face. So if
you think about your Saturday morning cartoons, those of you that are old enough, and some
— you know, something happens and, you know, the face just kind of accordions in like gnaaa
[phonetic sp] — kind of like that, right? And so that’s the kind of appearance you see
in the pug or the cane corso, and that’s very different than what you see in the Afghan
and the bull terrier, which have very elongated noses, and that’s referred to as a dolichocephalic
phenotype. So a long nose versus that very pushed-in face. So we’ve been going after
these kinds of genes, and we do the exact same experiment over and over, we comb the
genome, and we look for evidence of a selective sweep, and that’s what the data actually looks
like. So these are base pair positions along canine chromosome 30, and you see a certain
level of chatter, and then when you get right here, you can see this big dip. And that’s
telling us that there’s a selective sweep there, that that’s a place where there’s a
lot of homogeneity, that — something breeders have been selecting on for years and years
and years and years. So they don’t know that there’s a gene under there. They don’t know
what the gene is they’ve been selecting on, but we’re going to find it, and we’re going
to tell them what it is. Okay, so here are a set of 12 dog breeds that
my lab has now sequenced. We’ve sequenced these breeds pretty deeply so we’ve got a
pretty good genomic sequence, and we picked these breeds for lots and lots of reasons.
We have lots of different studies going on, and there’s another actually 40 dog breeds
that we and others have sequenced as well, but these were the first 12. And partly, we
picked them because if you think about going very brachycephalic to very dolichocephalic,
you have a really nice continuum. So we will be able to use this data to try and figure
out what’s underneath that little V, that statistical blip that the breeders have been
selecting on over and over by comparing the brachy- and dolichocephalic dogs. Now, you’ve heard a lot about the genome project,
and usually people talk about the human genome project. When I hear about the genome project,
I think about the canine genome project, because that’s what’s really important to me. So I
know this, again, looks like a checkerboard, but each one of the rows is giving us information
about the sequence variation we saw in those 12 dog breeds. So we started out with 190,000
base pairs, and 2,000 possible variants, and then we started filtering and filtering. We
got it down to 85,000 variants and 48 variants, and in the end, when we applied the sequence
of all those other breeds, we were able to find the single base pair in the single gene
that turns out to be important, and it’s how canine chromosome 30 contributes to that facial
phenotype. So the gene is called bone morphogenesis protein 3 — kind of makes sense it’s a bone
morphogenesis protein — and it is a single base pair change that changes one amino acid,
phenylalanine to lucy [phonetic sp]. One amino acid. Now, when Jeff came to me with that data,
I said, gosh, you know, I believe you, but in order to publish this, we’re, you know,
we’re going to have to have more proof. And so Jeff did lots of statistical studies, and
they all looked really, really good, and we wrote it up, and I said, you know, it’s pretty
good, but I think we’re just going to need just a little — a little bit more proof to
get it into one of the fancy journals. And Jeff said, “Well, okay. I can knock that gene
out in zebra fish, and I can make a pug-nose fish.” And I said, “You’re going to make a
pug-nose zebra fish?” And he said, “I’m going to make you a pug-nose zebra fish.” And so
there’s a technique he used, and he knocked that gene out, and that is exactly what Jeff
made. So let’s forget the top row for a second — I’ll
come back to that — but if you look where it says E, F, and G, that’s a zebra fish,
you know, several — about 48 hours after the embryo was fertilized, and it didn’t get
injected with the stuff that knocked out the gene. And then the blue is the cartilage from
the top of the jaw, the top jaw, and the G is the bottom jaw. And now, the other two
are examples of fish that did get injected with what we call morpholinos, and they knock
that gene out, and you can see that the jaw is gone, especially that bottom jaw. Look,
there’s almost no blue staining in J and M, and even the top jaw is pretty screwed up
as well. So in essence, Jeff made me a pug-nose zebra fish, and indeed, in doing so demonstrated
that by knocking out just that one gene, we’re able to dramatically affect the jaw structure.
And so this gene, in fact, does turn out to be important in one particular type of human
craniofacial abnormality. So this is another example of how, you know,
we started with healthy dogs, long-lived dogs, but they had a particular phenotype, this
sort of pushed in face, and we’ve been finding the genes responsible for that. And there’s
not just one; it turns out there’s actually several. And in aggregate, they are responsible
for the dramatic difference between being brachycephalic with a pushed in nose, or dolichocephalic
with that elongated nose. And we can prove we’re right by going to some of these model
organisms like zebra fish, which are these sort of little tiny fish that people use in
the lab sometimes. Okay, so those are examples of how my lab
has been studying morphology. And right about this point in time, someone usually says to
me, well, you know, that’s great, but — gosh, dogs have an awful lot of diseases. Do you
actually, directly study any of those diseases as well, and if you do, are they telling us
something about human disease? And the answer to both those questions is a resounding yes.
So here are the top 10 genetic diseases in dogs, and what do you notice is number one? Audience Response:
Cancer. Elaine Ostrander:
Right. About one in how many humans will get cancer in their lifetime? Anybody? About one
in four people will get cancer, some kind of cancer at some point in their life. They
may not die of it, but they’ll get it, and about one in three dogs will get cancer in
their lifetime as well. How many of you have had a dog or a cat who got cancer? Yeah, right.
So in my lab we do in fact study several different types of cancer. Now, this is one of my favorite
dog pictures ever. It was actually sent to me by a very well-known, and very generous
dog breeder, and she breeds — what are these? Audience Response:
Poodles. Elaine Ostrander:
Standard poodles. Right, these are standard poodles. So about four years ago, we started
getting phone calls from people who owned standard poodles, telling us that the dogs
were getting a particular kind of cancer, and it’s called squamous cell carcinoma. And
oddly, it was occurring in the toes, and sadly the way the veterinarians have to treat it
is they have to actually remove the toes. And so that’s horrible for the dogs, it’s
horrible for the owners. If the dog’s a show dog, it won’t be after that, and people obviously
don’t want to breed to those dogs after that, and so this is really a big deal for this
community, but what was so interesting about this is they said, you know, we only ever
see the disease in black standard poodles, and we never see it in white standard poodles.
And so we’ve now looked at hundreds of standard poodles, and we see it in black and brown,
very, very, very, very, dark gray, but we don’t see it in the white, or the cream, or
the apricot dogs. So we thought, well, you know, there’s a lot of selection for coat
color in standard poodles. Maybe they’ve inadvertently selecting for a cancer gene as well, and that
in fact turns out to be the case. Now, let me just tell you that we study lots
of kinds of cancer in my lab, and in fact, across the world dog geneticists study lots
of kind of cancer. So those of you who have long-limbed breeds like Scottish deer hounds
or Irish wolf hounds probably worry about osteosarcoma. We see tons of bladder cancer
in Scotties, and Westies, and Shelties, and we actually have a paper we’re writing about
that now that comes again from sequencing tumors, the DNA from tumors that we find in
those dogs. If you’ve got a Bernese mountain dog, a super wonderful dog breed that’s really
increasing in popularity, or a flat-coated retriever, one in four, one in six of those
dogs will get malignant histiocytosis or histiocytic sarcoma. Stomach cancer, we see in the Belgian
sheepdog, the Belgian Tervuren, as well as in the Chow Chow. Universally lethal; dogs
don’t survive it. And the idea here is we study these in dogs because the breed structure,
as I told you at the beginning, simplifies the overall problem. We’ll see shared genetics
among affected dogs, and it’ll be distinct from what we see of healthy dogs of the same
or, remembering the wheel, very related dog breeds. So this is a picture of squamous cell carcinoma.
You can see that the toe is sort of blown out. It’s the most common nail bed cancer
in dogs. If you are a giant schnauzer, your chances of getting this are 22-fold higher
than the average mixed breed dog walking down the street. If you are a Briard, your chances
are 10-fold higher than the average mixed breed dog walking down the street, and if
you’re a standard poodle, they’re about 6-fold higher, although standard poodles are where
we see most of this, because they’re the most popular of those three breeds. And again,
in the cases of the standard poodles, we only see it in the black dogs, not in the white.
For really complicated reasons in the Briard, we actually see it in black as well as white
dogs, and we figured out why that is. It’s actually a very, very complicated genetic
story, but let me tell you a little bit about standard poodles. So we, again, combed the genome with our 100,000
points of variation, and we found a signal on canine chromosome 15. It wasn’t as strong
as what we saw when we were looking at those morphologic traits, because breeders haven’t
been trying to breed cancer into dogs the way their trying to breed, you know, short
legs or large body size; but nevertheless, it’s there and it’s in a lot of dogs. Now,
I think this is the last data slides that I’ll show you. And so what we did is we found
a region on canine chromosome 15. It was about a million base pairs long. We sequenced it
in lots of affected and non-affected dogs, and everywhere that there was a possible mutation,
there is a triangle. And then, we defined a region that, you know, maybe if we were
liberal in our thinking, was about 500,000 bases in size; and if we’re conservative in
our thinking, it’s about 800,000 bases in size. And what’s cool is there’s only one
gene in that region, and that gene is called KIT ligand, and we knew instantly that we
had found the right gene, because it’s a gene that’s important in coat color, but it’s also
a gene that has been shown to be important in cancer. So we did a lot of work. It took
about three years, and we in fact, found the mutation, and in this case, we again found
sort of a new and interesting way that the genome gets screwed up. So it turns out that the mutation is, again,
extra DNA; it’s not a deletion. It’s an insertion of about 5,000 bases, and it can be present
one, two, three, four, five, or six times. And the more times it is present, the more
these green proteins bind; and the more these green proteins bind the more they ramp up
production of this gene KIT ligand; and the higher you ramp up production of this, the
greater the chances are that you’re going to get the cancer. So if you’re a dog who
has this insert present on both of your chromosomes four or more times, boy, your chances of getting
cancer are really, really high. If you have it maybe four times on one chromosome, three
on the other, it’s sort of moderate. But four is really the threshold. If both of your chromosomes
have it repeated three times or two times or one time, or any combination of thereof,
no chance you’re going to get cancer. No chance. And we’ve looked at hundreds and hundreds
of dogs, and we’ve never even found one. So this is one of these sort of threshold deals
where you have to get KIT ligand ramped up to a certain point in order for it to go ahead
and cause the cancer. So it makes it hard to develop a genetic predictive test, but
people are in the process of doing that. We found out why white standard poodles didn’t
get this. It isn’t because they didn’t carry the mutation, the four-four genotype. They
didn’t — they do indeed have chromosomes where this insert is present multiple times,
but they have a compensatory mutation on another gene called MC1R, and it completely knocks
this out. So it’s a case where they have a bad mutation, but they have a good mutation
and good triumphs over evil in this case, and so they never, ever get this, even though
they carry the bad genotype. So it’s an important lesson, because just looking for the presence
or absence of the bad genotype isn’t really totally predictive. You also have to look
for the presence or absence of the compensatory mutation, and all the white standard poodles
have it, none of the black standard poodles have it. And so that explains the difference
in what we observed in coat color. So I’m going to stop there. We have about
15 minutes to ask questions, or 12 minutes or so. I hope I’ve showed you that dogs are
a really fun system for looking at both simple and complex traits, including susceptibility
to cancer, which is of course a very complex trait. When we study morphology in dogs, we
learn things that are important about development of all mammals, and that includes humans as
well. For both canine and human health, these studies of cancer have become very, very important,
and there are labs really at vet schools, as well as at non-vet schools that are studying
every conceivable kind of cancer, as well as all the other common human diseases: epilepsy,
diabetes, heart disease — you know, whatever you’ve got including morphologic traits you
don’t like, like baldness or obesity or things like that. Those — there are labs all over
the world that are studying those things as well. Many of these studies are long term; some
are short term. There are a number of disease genes that we’ve been able to find the mutation
for. We’ve been able to develop a genetic test, and breeders are now using those genetic
tests to make really good decisions to produce healthier, more long-lived dogs. So you don’t
see kidney cancer in the German shepherd anymore, and we’ve been able to wipe out collie eye
anomaly, which is a degenerative disease of collies and border collies, and a number of
herding breeds, as well as several other diseases. And this has all been done in collaboration
with breeders and owners and veterinarians actually all over the world. Samples are always needed so if any of you
have a really interesting dog breed — I gave this talk yesterday, and a man came up to
me afterwards and said “I have a Shiba Inu if you want DNA from that.” Yes, we do want
DNA if you’ve got an interesting dog breed. We love your labs, we love your golden retrievers,
we love your German shepherds, but we probably have enough DNA on those, but the more esoteric
breeds, we’re still always collecting more DNA from. And you know, great progress can
be made, but you know, it is necessary to get the DNA samples, and allow us the time
to do our work. So for the breeders in the audience, I know many of you have contributed
samples, and you wonder how long it’s going to take. Wel, sometimes it’s a year, sometimes
it’s going to be five or six years, because some of these problems end up being simple;
some of them end up being very complex, but our goal is always to make available to you
some sort of a diagnostic test. So I’m going to go ahead and stop there, and
allow you to — turn up — maybe turn up the lights? Turn up the lights, and I’ll go ahead
and take questions. Okay? [applause] Elaine Ostrander:
Fire away. Female Speaker:
[inaudible] Elaine Ostrander:
This slide? How do we find out what? Female Speaker:
How do you find out the disease risk? Elaine Ostrander:
How do we find out the disease risk? So what we did is we got DNA from a lot of poodles
who had the disease and a lot of poodles who didn’t have the disease, and then we sequenced
it to see how many times this was re-iterated, how many times this particular 5.7 base pair
unit was repeated. And we actually knew from looking at the human genome, we knew exactly
what these 5,000 base pairs do. We knew that this protein binds to them, and when it does
it ramps up production of KIT ligand. So we can do statistics looking at dogs who have
two copies, three copies, four copies, five copies, and we can look at them at 10 years,
11 years, 12 years, 13 years, 14 years, and see who does and doesn’t have cancer. And
from that, we can figure out what the risk is for a dog who has three copies or four
copies or five copies, and four is really the cutoff. If both of your chromosomes have
three copies or two copies, or one copy, we’ve looked at hundreds of dogs, and not a one
of those dogs has this kind of cancer, but as soon as one chromosome gets four cancer
— has the repeat — repeated four times, then we start to see the incidents of cancer
creeping up, and the more you have, the higher it gets. Okay? And there are you know formal
statistical tests that we can apply to that, and if you’re interested, if sort of got a
statistical mind, come see me afterwards and I’ll give you the paper and the tests that
we use. Okay, other questions? Yep? Female Speaker:
I notice that you were [inaudible] dogs [inaudible]? Elaine Ostrander:
Sure. So the question was, is epilepsy indeed prominent in dogs, and yes, we see it in lots
and lots of dog breeds; and have we found genes? We’ve been part of one study that identified
one gene, published it with a group in Belgium several years ago in Science, but lots of
other people are looking at this problem, because you know, not only is it an important
problem in human, it’s a big deal in dogs. I mean, if your dog has epilepsy, I mean,
that is a lifelong problem that you as a pet owner have to deal with, and certainly those
dogs don’t show and people don’t, you know, put them in the breeding pool if they know.
And so some of those genes have been found; not all of them have been found. And there
are some breeds where it is a much worse problem than it is in other breeds. So that’s one
of the areas where we see some of the most active and intense work. Yeah, you had a question. Male Speaker:
This is sort of a general [unintelligible] not so much a question. Elaine Ostrander:
Okay, I probably won’t know the answer, but go ahead. Male Speaker:
Well, I was just wondering what sort of breeds made up the dog reference gene? Elaine Ostrander:
Oh yeah, sure I do, because I picked the breed for the reference genome. So he asked me,
what breed or breeds make up the dog reference genome? So it’s one breed, and it’s one single
dog. So in 2001 I had the chance to pick the dog that was going to be sequenced, and you
know, truthfully what I did is I probably looked at 100 different dogs of all breeds,
and I picked the most in-breed dog I could find. And the reason for that is the way genomes
are sequenced is not from the top of chromosome 1 to the bottom of chromosome 38. What they
do is they cut the DNA up into a zillion little pieces, and then they sequence it all randomly,
and then they have a computer put it back together. So if what you got from mom is really
different than what you got from dad, it’s a harder computational problem. If what you
got from mom and dad was pretty similar, it a much simpler computational problem. So I tested 100 dogs at 100 places in their
genome, and there was one dog that was easily the most in-breed. It was a boxer, and it
was from New York state, and as luck would have it, it was actually owned by a veterinarian.
So when I went to him and I explained that his dog was the lucky winner — I mean, he
actually understood that this was really important. He provided us with an awful lot of DNA, and
he understood that, you know, when the dog — it was a pet, you know, like a lot of our
dogs are — that when the dog died that we, you know, we would like to get some samples
from that dog, because it was, in fact, going to be the reference genome. And his only request
that I not divulge his name, which I’ve never done, or his location, which I’ve never done.
So he was great, he was fantastic, but it was a boxer, and her — it was a she. Almost
all the reference genomes, except maybe maybe — are a she, because then we get good data
on the X chromosome. Female Speaker:
Have you guys found a gene related to like a Yorkie, just randomly like [inuadible]? Elaine Ostrander:
So is — have we found a gene linked to fur loss in breeds like the Yorkie? So I haven’t
looked at that. There are breeds, like the Chinese crested or the Mexican hairless, there
are breeds that have very little fur, except for some tufts at the top of their head and
down by their toes, and those genes have in fact been found, but those dogs, that’s part
of the breed standard; that’s how they’re supposed to look. When a dog blows all of
its fur, I mean — and that’s an anomalous sort of thing — I’m sure there are people
looking at it. I’m not one of them, and I’m not aware of a paper, but I could certainly
tell where in the literature to look for it. Yeah? Male Speaker:
Are you able to define what would be the top, like healthiest breeds? Elaine Ostrander:
Sure. Yeah, so the question is what are the healthiest breeds, and it’s a little bit of
a trick question, and the reason is because within every breed there are lines that are
really healthy, the breeders are very savvy, they’ve gotten dogs from multiple places in
the world that are members of that breed, or multiple places in the United States that
are members of that breed, and they’ve worked really hard to maintain the hybrid rigor of
that breed; and there are other breeders, you know, who have bred a lot of closely-related
individuals one to another, and their lines may look good, but they have a lot of health
problems. So there’s no one right answer. I mean, you can look on the internet and,
you know, they’ll certainly give you those — that kind of information, and they’ll say
things like, well, you see a lot of cancer in boxers or in golden retrievers; or you
see a lot of copper toxicosis in the Bedlington terrier. You see — you know, there’s different
diseases that tend to predominate in different breeds. There are some situations where disease
is so predominate, like copper toxicosis was in the Bedlington, before the gene was found
that I think that was a fair comment. You know, there are other cases where I think
you just have to really search. I mean, I wouldn’t be hesitant to own a golden retriever
or you know, a German shepherd, or any of these other really popular breeds. I would
just work really hard to talk to people the breeder had sold to, and find dogs that lived
a really long — a really long life. In general, small dogs live longer for sure
— the terriers, the toy dogs — in general, they do live longer. And dogs that have a
working job like border collies, boy, some of those live 17, 18 years. My border collie
lived to be 13 years. So breeds where there’s a working lineage — I mean, they have to
healthy to work — those tend to be pretty long-lived. The really big breeds tend to
have more heart difficulties and problems — Saint Bernards, and you know, some of these
giant breeds where they’ve just almost been bred to be too big for their heart. You know,
they rarely live beyond the age of 10 or 11, because they almost always go for — from
heart problems. So, you know, I always tell people to, you know, pick the breed you want,
pick the breed that matches your family. There’s all kinds of tests on the internet to help
you do that. But then work with — you know, really look around and take your time to find
a healthy and reliable breeder. Yeah? Female Speaker:
Would you say that breeds that are closer to the wolf, like in ancestral [inaudible]
lineage, would you say that they are [inaudible]? Elaine Ostrander:
No. So she — the question she asked is, are breeds that are more related to wolf healthier,
and I guess I would say no, you know, in part because things that maybe look more wolfish,
like the Malamute or the husky, I mean, you know, there were clearly multiple domestication
events that occurred in multiple places in the world, and you know, by and large, truly
it’s the small dogs that are the healthiest and really do live the longest, and so I wouldn’t
necessarily say those are, you know, among the healthiest, although, you know, I had
a Malamute that lived to be like 15 or 16 so, you know, again dogs that have a working
function, but this was bought from a line of dogs that, you know, were involved in mushing,
and so that’s — you know, those are going to be healthy dogs so I wouldn’t necessarily
say that. Other questions? Yeah? Female Speaker:
Do mixed breeds do better than pure breeds? Elaine Ostrander:
Do mixed breeds do better than pure breeds? You know, everybody thinks that they’re going
to, and I have so many people come up to me and say “I bought a Labradoodle or a” — you
know, they have all these weird, cool names, and they said, you know — and so — hybrid,
bigger, you know, I’m — my dog’s going to live to be 22, and then they’re shocked when
they don’t. And the reason is because, you know, if the parent breeds or the parent lines
were themselves not healthy, particularly if they were not healthy because they had
the same disease or the same mutation, you haven’t — it doesn’t do any good, right?
And so you know golden retrievers, you know, they get a bad reputation for having a lot
of inherited disease. I think there are some lines of golden retrievers that are wonderful
out there, but there are also some lines that really do have a lot of disease, and because
they’re so good with families, they’re involved in an awful lot of crosses, and you know,
you can say that of lots and lots of other breeds. So, you know, in general, what I like
about pure bred dogs is you sort of know what you’re getting. You know, most poodles I know
live long, long, long, long periods of time. There an awesome breed in all three sizes.
They have a reputation in the breed club for being very vigilant, being very careful, and
so I don’t necessarily think that’s really the best choice. Yeah? Female Speaker:
[unintelligible] you talking about being able to remove the mutation? So is that happening
widely within that breed — Elaine Ostrander:
So — Female Speaker:
— or is there some special [unintelligible]? Elain Ostrander:
— the question is, you know, removing mutations, does that happen widely? So it really isn’t
removing mutations in the sense of how that language is used, but it’s really about breeding
it out. So breeding a carrier not to another carrier, but to a healthy dog. And when I
talk to breed clubs, I tell them first thing, don’t throw all the carriers out of the breeding
pool, because they’re contributing so many good things that you’re going to just end
up with something else you didn’t have a problem with before. But — but take your carrier
and breed him to a non-carrier. The carrier — get the progeny tested, breed carriers
to non-carriers, and gradually breed it out. And we’ve seen examples where a small breed
has thrown out all the carriers, and they have a disease that’s really prevalent, and
then suddenly they have three other diseases that crop up as recessive. The American Kennel Club has just been wonderful.
I mean, they ask us to come to all their meetings and their specialty events, and their trials,
and we are inundated with invitations to come and give talks about exactly these kinds of
issues, and labs like mine often make the data available without patenting it. Sometimes
it is patented, but often without patenting it, just so the tests can get out there and
people can start using it. And breeders are some of the smartest geneticists in the world.
They absolutely use it, because it is their livelihood. They want to breed healthier,
more long-lived dogs, and if they’re the first off the block with a reputation for using
genomics to breed healthier more long-lived dogs, people love that — people love that.
And so I have found that this — I mean, I have 50,000 DNA samples in my freezer, and
I can think of one incidence where I asked for a DNA sample and I was turned down. Go
ahead. Male Speaker:
My question is back to the mixing question. So in agriculture, the way you find new traits
[unintelligible] is by crossing breeds so you will get the non-additive effects and
things like that. Is there a push in the dog world to try these things out and see what
happens by mixing different breeds? Elaine Ostrander:
No. No. There isn’t. Male Speaker:
[unintelligible]? Elaine Ostrander:
Right. So the question that was asked was, is there a push in the dog world to, you know,
mix dogs of different breeds to try and figure out what’s going on with the genetics of some
of these traits? I just put this up because it’s my favorite picture in the slide show.
And the answer to that is no. First of all, we don’t breed or keep any dogs. So we’re
not going to do that. And in the pure bred dog world, you know, people — you know the
convention is you breed dogs of one breed only to members of the same breed, and that’s
just not the convention in that community, you know, to take a dog that may be a popular
sire and breed him to a bunch of other breeds to figure out what’s going on. That’s just
not what is done. So it really isn’t. And you know, I’m not in the management of the
American Kennel Club so it’s not really for me to say, but — okay, one more? Can take
one more question? All right. One more question. Sure. Male Speaker:
You mentioned about 50,000 data points. Are they all pure breed or do you also have mixed
breeds and mutts? Elaine Ostrander:
So I take 100,000 data points, and I have 50,000 DNA samples from dogs in my freezer.
I have — my lab has very few mixed breed dogs. Almost all of those are from pure bred
dogs. They’re not all American Kennel Club recognized breeds. You know, some of them
are odd, you know, European or Asian breeds. They’re not all AKC breeds, but they are pure
breeds; they’re not mixed breeds. But there are other labs that have focused their studies
on mixed breeds. So there’s one at Cornell who specializes in village dogs, and he’s
traveled all over Mexico, South America, Africa, to the outskirts of town on the garbage dumps,
and he and his team sample hundreds and hundreds and hundreds of mixed breed dogs, and that’s,
you know, his thing. So, you know, you kind of can’t do everything, and so that’s been,
you know, where my focus has been, but there are certainly people doing that. All right,
I’m going to let you go. There’s lots to see out in the museum, and I’ll be around for
questions, and thank you all for your attention.

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1 Response

  1. Reto Fassbind says:

    Excellent, easy to understand and well presented and instructive insight into the genetic study of different dogs breads and their traits and diseases.

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