Investigation of Genes Controlling Pigmentation in Alpacas
Alpaca Research Foundation (ARF) Investigator Profile: David L. Kooyman, PhD
By Ingrid Wood (Stormwind Alpacas)
During my long teaching career, I once taught a class of fifth graders about the
physiological similarities shared by human and pigs. Body parts from pigs can be
used to replace, for example, defective heart valves in humans. “She’s crazy,”
one of my students whispered, sotto voce. Several heads nodded in silent
agreement. Too bad my students did not have the opportunity to talk to David
L.Kooyman, PhD, a scientist presently working in the Department of Physiology
and Developmental Biology at Brigham Young University in Utah. His biography
profile includes employment at Baxter International in New Jersey. As a senior
scientist, Dr. Kooyman’s worked on xenotransplantation, a project where he
helped to clone genes and genetically engineer pigs to be used as organ donors
for humans.
Pigs were not Dr. Kooyman’s first research interest as far as species go. While
working on a Masters Degree at California State Polytechnic University, his
attention was focused on the reproductive physiology of goats. After completing
a PhD in molecular and cellular biology at Ohio University in Athens and a post
doc at DNX in Princeton, Dr. Kooyman eventually worked for previously mentioned
Baxter International. His academic achievements and interests finally led to his
present position at Brigham Young University. “I served several years as
Associate Dean in the College of Biology and Agriculture,” Dr. Kooyman
mentioned. “Prior to that, I was Chair of the Animal Science Department. I
resigned my position as Associate Dean because I wanted to return to my first
love… teaching and research.”
“How does the alpaca project fit into your schedule?” I wondered, assuming that
camelid research played only a very minor role among Dr. Kooyman’s multitude of
academic duties. His answer surprised and delighted me. “My research focus now
is identifying and characterizing economically important genetic traits in South
American Camelids,”
he replied. His lab at Brigham Young University is the first and possibly the
only lab in the world that produces llama bacterial artificial chromosome (BAC)
“libraries”. In a BAC library, large chunks of llama DNA are cloned and
placed within a man-made, artificial chromosome in bacteria. “You are continuing
the research begun by Dr. Emily Campbell?” I asked. “Yes, we are in the process
of identifying and describing the genes associated with color in llamas and
alpacas,” Dr. Kooyman confirmed. I could hear the happy satisfaction in
his voice when he added, “We’ve made quite a bit of progress in our research.”
The interview and subsequent communication with Dr. Kooyman reminded me once
again how alpaca breeders have been fortunate to “piggy –back” on the results of
research conducted on other species. Dr. Kooyman wrote, “We have found that
identifying candidate genes for studying alpaca fleece color is possible because
of the high conservation of these genes in other domestic animals previously
studied and characterized.” He specifically mentioned pigs, cattle, and horses.
Alpaca breeders need to appreciate the fact that the inheritance of color/
patterns is the result of bio-chemical processes programmed by genes which code
for pigment or loss thereof. Some breeding results seem bizarre to those who
have never studied this science. For example, two black alpacas may produce a
white cria while, on a neighboring farm, two white alpacas are the parents of a
black infant. Does this make sense? It does! An animal’s final color phenotype
is the result of many genes. The white alpacas in our examples are, genetically
speaking, black alpacas. Their pigment was “stripped” by various genes coding
for lack of pigment. The mating of sire and dam caused a series of genes to be
“re-shuffled”, with obviously eye-popping results.
Let’s remember that only two pigments exist in mammals: black (eumelanin) and
red (pheomelanin – sometimes spelled phaeomelanin). Two major genetic “players”
determine which pigment (s) are produced by the pigment producing cells, called
melanocytes. These genes are found at two loci (genetic “addresses”): the Agouti
locus and the Extension locus. A large supporting cast of other genes code for
shading/no shading, dilution/no dilution, and pattern/no pattern. Regardless of
these modifications, we must think of all alpacas as either red or black. To use
several examples: a light fawn alpaca is red, a silver grey is black, a medium
brown is red, a rose grey is red, a fawn pinto is red and so on. In terms of Dr.
Kooyman’s present research project, we need to primarily concern ourselves with
those genes found at the Extension locus. However, we can’t get around
discussing genetic mechanisms found at the Agouti locus. Shortly, you will see
why that is necessary.
I mentioned that the function of genes at the Agouti and Extension loci has been
firmly established in other species. Dr. Kooyman, along with other scientists
such as Dr. Phillip Sponenberg, hypothesize that the Melancortin 1 Receptor
(MC1R) gene at the Extension locus and the Agouti Signaling Inhibitor Protein (ASIP)
gene at the Agouti locus play important roles in the differentiation of red
versus black phenotypes in alpacas as well. The close bio-chemical interaction
between the products of these two genes (MC1R and ASIP) can be very complicated.
Dr. Kooyman patiently explained how the melanocyte (the body cell that produces
pigment) constantly produces pheomelanin (red) if it doesn’t get the directive
to produce eumelanin (black). “Tell me what happens, bio-chemically speaking, to
make mammals black?” I asked. “In that case, a hormone called Melanocyte
Stimulating Hormone (MSH) is involved,” Dr. Kooyman answered. He emphasized, “If
MSH is bound to its receptor MC1R, the pigment cell will be stimulated to
convert pheomelanin into eumelanin and the animal will be black.”
Looking over my notes several weeks after the interview, I thought, “Now what
exactly does “bound to” mean?” Minutes later, I was on the phone to Dr. Patricia
Craven, a fellow alpaca breeder (Cherry Ridge Alpacas), ARF board member, and a
friend. She has the patience of a saint when I call on her knowledge as a
research scientist to explain scientific concepts to me. “Yes, I can fill you in
a little more,” Pat responded to my plea for help. “MSH is a peptide hormone and
is secreted into the blood stream from the pituitary gland. MC1R is a receptor
that is located on the cell membrane, the outer layer of the cell. MSH doesn’t
enter the cell.” “Then how does this hormone do its job?” I interrupted. “MSH
interacts with its receptor, MC1R, in the cell membrane,” Pat continued,
unruffled. “The receptor recognizes the hormone and transmits a signal from the
hormone to the cell. The signal causes the enzymatic conversion of pheomelanin
to eumelanin.” Well, who would have thought that hormones play a part in color
genetics?
During the interview, I had questioned Dr. Kooyman at this point, “But not all
mammals are black, so MSH (the hormone) either isn’t produced or doesn’t work
all the time?” That’s correct”, he replied. “When ASIP is bound to MC1R, MSH is
unable to bind as well and only pheomelanin (red) will be produced. The actual
color will depend on the other color determining genes of the animal. In order
to understand how this occurs, it’s necessary to learn a somewhat complicated
genetic concept. Some genes have mutated variations of the original form. We
call those alleles.” Dr. Kooyman sounded almost apologetic as he added
diplomatically, “That’s very hard to understand. I can’t think of an easy way to
explain it.”
“You can think of alleles as ice-cream flavors…,” I started my somewhat goofy
definition of an allele in layman’s terms. Dr. Kooyman laughed. “Yes, that’s a
good analogy,” he acknowledged.
So, let’s talk ice cream. It comes in flavors such as vanilla and “mutated”
versions such as chocolate, strawberry, lemon, and mocca. A person has only two
hands and can only carry one cone (one flavor) per hand. They are all ice-cream
(gene) but different flavors (alleles). Likewise, at each locus, an animal
carries only two genes – one inherited from the sire, the other one from the
dam. If the genetic material on that locus is the same for the entire population
(ice-cream analogy: vanilla-vanilla or mocca-mocca to use two examples),
scientists use the word gene. If the flavors (alleles) vary within a population
(examples: vanilla-strawberry, chocolate-vanilla, mocca-mocca, strawberry-mocca),
the variations are referred to as alleles. Combinations can vary considerably
within a specific population, but alleles can also be easily lost forever due to
natural or artificial (human) selection pressure. (A friendly warning to alpaca
breeders: think long and hard before you call for specific colors/patterns to be
eliminated from the entire North-American alpaca population!).
In any case, scientists established the existence of multiple alleles of the
MC1R gene in humans, dogs, cattle, pigs, horses, goats, and probably many
others. In many mammals, multiple allelism also occurs in the gene coding for
Agouti Signaling Inhibitor Protein (ASIP). So how does the potential for
multiple alleles at the Extension or Agouti locus play a role in determining
whether an animal is black or red? Let me give four examples based on studies in
other species.
1. The animal could have inherited an allele at the Extension locus that codes
for a form of the MC1R that does not bind MSH and is therefore not functional.
Dr. Kooyman explained the bio-chemical process, “The non-functional MC1R allele
produces a red phenotype because the hormone (MSH) is not able to bind to its
receptor, MC1R.” He further mentioned that the mutated MC1R allele coding for
loss of function can produce a wide range of “red” phenotypes – from a rich,
dark red to fawn so light that it appears white. He pointed to the Black Bear as
an example of this phenomenon.
2. The animal could have inherited a functional allele of the MC1R gene at the
Extension locus plus a functional allele of ASIP at the Agouti locus. A
functioning ASIP allele would lead to production of ASIP, an inhibitor protein
that prevents MSH binding to its receptor. This animal would also be red.
3. The animal could have inherited a functional allele of the MC1R gene at the
Extension locus plus two non functioning alleles at the Agouti locus. In the
latter case ASIP would not be produced and therefore could not interfere with
the binding of MSH to its receptor, MC1R. This animal would be black.
4. As with just about everything in science (or so it seems to me), there are
exceptions to the rule. Dr. Kooyman made it clear that, in some animals, a
dominant black MC1R allele produces a receptor that allows the hormone MSH to
bind even if the ASIP allele is functional and ASIP is produced.
It is quite possible that alpacas, like dogs, have more than two Agouti locus
alleles (flavors) to “choose” from. Once again, let’s remember that the
individual alpaca carries only two alleles at each locus. After examining
breeding records, Dr. Sponenberg in fact proposed a series of Agouti alleles,
and their identification certainly lends itself to another project funded by the
Alpaca Research Foundation (www.alpacaresearchfoundation.org).
At this time, let’s clearly define Dr. Kooyman’s research objective: in his
laboratory at Brigham Young University, he will identify all the alleles of the
MC1R gene in alpacas within the available population. “Comparing genotypes to
phenotypes will help to determine if multiple alleles exist at the MC1R gene in
alpacas,” Dr. Kooyman further clarified his mission.
The interview drifted back to more private matters. Dr. Kooyman’s interest in
camelids developed in 2002. His oldest daughter Kristy owned llamas and alpacas
at that time.
She urged her father to apply his considerable knowledge and expertise to the
animals she loved and Dr. Kooyman described as “delightful”. In 2004, he made
contact with South American scientists involved in camelid research. This led to
his election as lead scientist of the “Camelid Focus Group”. The group’s goal is
a comprehensive, integrated project in camelid production with the initial
emphasis on fleece characteristics including color and elimination of guard
hair. Dr. Kooyman expressed a keen interest in helping the people of the Alto
Plano. “Many live in poverty,” he stated, “my contribution to their welfare can
be in the area of genetics.”
Dr. Kooyman shared with me that he and his wife have four children, one of which
died a little over a year ago. When, at some time during the conversation, I
referred to his three children, he quietly but firmly corrected me, “No, we have
four children.” This loving inclusion of the deceased child in such a natural,
unaffected manner made a deep impression on me.
We talked about Dr. Kooyman’s upbringing on the edge of a rural environment in
Iowa. Summers were spent on his uncle’s farm helping to raise crops,
cattle, and pigs. Ah yes, those pigs of organ donor fame! It seems like quite a
journey from feeding slop to pigs to researching color genetics in camelids. It
is an equal stretch for many alpaca breeders, most of whom are not scientists,
to comprehend the complexity of color inheritance. The knowledge of bio-chemical
processes is crucial for a truly deeper understanding of the subject area.
However, it is not, in my opinion, a prerequisite for learning what I call the
“system” of color genetics.
Long before the first alpaca stepped on North American soil, scientists and
breeders developed a system of letters to express the mechanisms responsible for
mammalian color phenotypes. It also enhances communication among breeders. An
article describing the various color loci and genotypes of fancy mice is
entirely comprehensible to a breeder of dogs and horses. When Dr. Kooyman is
ready to share his research results, I will present them in this “breeders’
language”. The words Melanocortin 1 Receptor gene need not even be mentioned.
Breeders will be able to take Dr. Kooyman’s findings and apply them to their
breeding programs. This is indeed an exciting project and only the mere
beginning of what a scientist with Dr. Kooyman’s knowledge can accomplish for
the benefit of the alpaca community.
Ingrid Wood is the owner of Stormwind Alpacas. Her small farm is located in a
rural community in New Jersey. She offers a PowerPoint presentation An
Introduction to Camelid Color Genes to interested groups and private parties.
Her website,
www.StormwindAlpacas.com includes
all contact information.