University
of Cincinnati research shows that thyroid hormone receptor alpha plays
an important role in hind limb development in frogs. The results of this
study may shed light on the importance of hormones in early development
in humans.
University
of Cincinnati endocrinology researchers were recently able to mutate
the thyroid hormone receptor (THR) in one of two cells during the first
step of early egg division in tadpoles. As a result, they have
successfully disrupted the developmental timing of the hind limbs,
showing clear evidence for the importance of THR in the early
development of vertebrates. The results of this study may also have the
potential to shed light on the importance of hormones in early
development in humans.
With
new gene mutation technology developed in the last two years, UC
researcher Daniel Buchholz, associate professor of biological sciences
and graduate student Jinyoung Choi, along with scientists in the
Department of Mathematical and Life Sciences at Hiroshima University,
were able to successfully mutate the gene in the tadpole models.
Together, they found the value of tadpoles as ideal models for studying
the role of hormones in development because of the timely metamorphosis
from tadpole to juvenile frog, and because that transition is completely
dependent on hormones.
In
earlier studies, Buchholz found that tadpoles don’t metamorphose in the
absence of hormones. They instead just become larger tadpoles.
Hind
limb phenotype in F1 offspring. Representative sibling offspring from a
pair of TR! TALEN founders were imaged at feeding stage (upper panel).
The developmental difference in hind limbs is shown in the lower panels.
The hind limbs are bracketed.
This
phenomenon was first discovered in 1916 when scientists were able to
surgically remove the thyroid gland and found out that thyroid hormone
is required for metamorphosis. Now, at the almost 100-year anniversary
of this revelation, Buchholz and Choi are now able to study the other
part of the story.
“Now
we can manipulate the genes and the proteins that are the receptors to
look even further into what these receptors do,” says Buchholz. “This
new technology has been cited in Science and Nature Magazines and could
very well revolutionize the study of non-model organisms. Other
scientists have been using this technology in other organisms, but we
are one of the first to use the technology in tadpoles.”
Since
Hiroshima University was already using this new technique, they
supplied Buchholz and Choi with the technology and sent the reagents
here to make the mutation.
During
the first step of development where the egg divides into two, Choi was
able to mutate the THR in only one of the cells. While that cell makes
up half the body, Choi was able to label what cell she manipulated,
which gave rise to that half of the body. From that she could determine
which side had the mutation and which side was normal. They then looked
at what happened during development and consequently had a perfect
control inside the same animal.
Buchholz
explained that the study gets even more remarkable. Ironically, his
post-doc advisor Yun-Bo Shi at the National Institutes of Child Health
and Development (NICHD) –– where Buchholz first looked at molecular
biology in frogs from 2000 to 2006 –– also studied this exact technology
on tadpole models at the same time. While Buchholz, Choi nor Shi at
NICHD knew about each others' studies until recently, they all found the
same result after mutating only one hormone receptor.
In
a sort of twin study, UC and NICHD replicated each others' studies and
found identical results. As a consequence, both papers are published in
Endocrinology at the same time and the publication will also produce a
News and Views article on this topic because of the unique situation.
In
mice, Buchholz points out that scientists have been able to knock out
the gene for quite awhile because of the special structure of their
reproduction system that can make embryonic stem cells. But in tadpoles,
Choi and Buchholz were able to remove just one of the two (alpha and
beta) thyroid hormone receptors. By knocking out only the
alpha-receptor, Choi was better able to determine what each receptor
controls in an organism.
“We
already know what will happen if there is no thyroid hormone signal, as
it will simply take away the hormone,” says Buchholz. “In humans, no
hormone at all creates cretinism where the person has short stature and
mental retardation. So in that case it becomes quite severe.”
In
humans, Buchholz also explains that THR alpha controls heart rate, and
THR beta controls thyroid hormone levels, and during development, also
controls hearing. So being able to distinguish what one receptor does to
the other is pretty important and has distinct consequences, especially
when compared to just no hormone at all.
Frog
metamorphosis has been compared to birth in humans because during frog
metamorphosis there is a peak in blood levels of thyroid hormone
dependence, and there is also a peak in thyroid hormone blood levels at
the moment of birth in humans. Since humans and frogs are both
vertebrates, Buchholz explains that there are many other similarities
and the thyroid hormone receptors alpha and beta are expressed in
similar cells types.
The
cells can respond to similar agonists and antagonists, which are
chemicals that can block or induce thyroid hormone function. With this
technology, we can test those kinds of chemicals and study what effects
they have on the role of the receptors.
“Knowing
that what the THRs do in frogs is very similar to what they do in
people, we can hopefully better understand what is happening in people
during the developmental stage, which is very difficult to study in
humans,” says Buchholz.
Other Contact Phone: (513) 556-1824/Photos By: Daniel Buchholz
Funding: Junyoung Choi earned a $3,000. Sigma Chi grant in aid of this research.
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