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A few of the molecular components involved in mechanosensory
perception and implicated in deafness, however, have been
identifiedboth by Mueller and in other laboratories.
And Mueller and his laboratory are looking to find more
of these components by developing models of deafness, looking
for important proteins, and setting up in vitro and
in vivo systems to test the effects of those proteins
on mechanosensory pathways. They would also like to find possible
therapeutic targets among these components.
The odds of achieving this goal are helped by the fact that
the field has lots of good in vivo models that copy the disease
phenotype and can be used for finding the genes that cause
the disease. "This is fortunate," says Mueller. "A lot of
human diseases are so complex that you can't make decent models."
Many of the models of deafness have been around for years,
and some of them have names like jerker, waltzer, and shaker,
which reflect phenotypes arising from inner ear defects and
balance problems. Several other models have been created by
Mueller and his colleagues.
These models can be used to "positionally clone" the genes
that are responsible for the phenotype. Positional cloning
traditionally entails the use of classical genetic mapping
methods to confine the location of the gene to a particular
area in the genome, extensive sequencing of the region in
question, and the performance of computer-aided searches through
databases to find homology between sequences in that region
and known genes.
This has been a fruitful approach so far, and to date mutations
in at least 50 genes that code for everything from cell surface
receptors to soluble cytosolic proteins have been cloned through
these methods and were found to contribute to deafness. These
include five genes that lead to different forms of Usher disease.
Recently, Mueller and his laboratory published a paper in
the Proceedings of the National Academy of Sciences showing
that two of these Usher genes act in a common pathway and
in physical contact with one another.
One is a receptor protein cadherin 23. "We think that this
receptor connects the stereocilia and is intimately engaged
in the mechanical perception process by regulating the mechanical
properties of the cell," says Mueller.
This is an important role, because in the inner ear, stereocilia
appear in symmetric arrays and have to be connected to each
other at the tips, sides, and ankles to move as one unit and
properly detect sound waves. Cadherin 23 appears to be one
of the molecules that makes these connections.
Significantly, cadherin 23 also contacts the protein harmoninboth
are expressed in the stereocilia of hair cells. Mueller and
his colleagues found that cadherin 23 and harmonin interact
and bind to each other through particular "PDZ domains" in
the harmonin proteins.
This interaction seems to be necessary for hearing, and
when the two proteins do not bind to each other, the integrity
of the stereocilia is compromised. Mutations in either the
cadherin 23 receptor or the PDZ domain of the harmonin protein
can lead to Usher syndrome. Instead of normal, symmetrically
arrayed stereocilia, the mutations cause the stereocilia to
be splayed and unable to properly detect sound waves.
Also interesting is the finding that Mueller and his laboratory
made that the cadherin 23 receptor appears to be alternatively
spliced in the ear and in the eye.
"We think that you express two different complexes in the
different tissues," says Mueller. "One that is optimized for
mechanical sensation, and the other one [we think] somehow
has to do with maintaining the photoreceptor layer."
This may explain why defects in the gene encoding the receptor
protein lead to deafness at birth but contribute to blindness
that is only fully manifested later in life, when the mechanosensors
in the eye reach maturity.
Integrins and Neuronal Development
Another part of Mueller's laboratory is devoted to looking
at developing stem cells and the mechanics of differentiation
and migrationtwo of the most important processes in
During fetal development a fountainhead of brain cells,
hundreds of millions of them, are produced through proliferation
of stem cells. But at some point, these same cells have to
stop dividing, differentiate, and migrate to the position
where they exert their function as specialized neuronal cells.
Peripheral nerve cells also have to migrate to reach the area
in the body where they exert their function. While migrating,
these cells interact with other cells, which act as guideposts
along the way.
"What we're trying to understand," he says, "is how a stem
cell or a committed neuronal precursor decides to acquire
a particular differentiated state and migrate to a particular
When a cell changes from proliferation to differentiation
into a specialized cell, there are any number of gross and
subtle changes in the expression of genes within that cell.
Mueller and his laboratory look at expression profiles related
to the differential state of the cells, trying to get at the
molecular machines that regulate the switch from a proliferating
cell to a migrating, differentiating neuron.
They are also interested in how the mutations in the genes
they identify might contribute to pathology of central nervous
system diseases. Once he finds genes that he thinks will be
interesting to study further, he looks to perturb that gene
and probe its function by knocking out genes expressed in
the central nervous system through a sophisticated tissue-specific
gene targeting technology.
One of the genes that the lab recently knocked out in the
central nervous system (CNS) is the integrin subunit beta-1,
which dimerizes with a number of different alpha subunits
to form a functional integrin. Integrins are what cells use
to cling to the extracellular matrix.
Knocking out the integrins in the entire organism is not
possible because these adhesion molecules are necessary for
maintaining the integrity of tissue. However, knocking them
out specifically in the CNS creates a model with defects in
the neuronal glial cells that resembles a set of human conditions
known as lissencephaly, which comes from the Greek root for
This model displays phenotypes that resemble those of a
number of rare human diseases as well, such as Walker-Warburg
Syndrome, muscle-eye-brain disease, and Baraitser-Winter syndrome.
All of these diseases have common features, including smoothness
of the brain and defects in the brain's basal membrane. Patients
with these diseases also suffer, to a certain degree, from
retinal abnormalities, peripheral neuropahy, and muscular
dystrophy. Similarly, patients with non-lissencephalous forms
of muscular dystrophy sometimes have impaired mental function.
"What this all points to is a common molecular pathway for
these different human diseases that involves interactions
of cells with the extracellular matrix," says Mueller. "All
these diseases are not so unrelated."
A Working LaboratoryAlmost
Coming to TSRI and the ICND was a homecoming of sorts for
Mueller. He spent the majority of his graduate years in the
United States, working in the Lewis Thomas Laboratory at Princeton
University, and completing his postdoctoral work at the University
of California, San Francisco.
The move has not been simple. Setting up any new laboratory
is difficult, but moving an existing laboratory from a European
country to the United States offers its own unique challenges.
Much of his time in the last year has been occupied working
with the U.S. Department of Agriculture to get hundreds of
serum samples, antibodies, and other biological compounds
documented and approved for shipping and arranging visas for
his group. Mueller arrived with three graduate students, one
laboratory technician (now manager), and two postdoctoral
fellowsa few of whom came a little early to start making
Now, near the end of this long process, he is looking forward
rather than back.
"March was the deadline I set to start to do experiments,"
says Mueller. "We're almost theretoday we start doing
little things, and next week, we're on the bench."
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