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The Skaggs Institute
for Chemical Biology
Fundamental Processes in Neural Development
G.M. Edelman, A. Atkins, S. Chappell,
J. Dresios, O. Harismendy, K.N. Gonzalez, D.C.Y. Koh, P. Panopoulos, J.
Pilotte, M. Tsatmali
We continue to
delineate primary cellular processes and their role in the development of the vertebrate
nervous system. In the past few years, this effort has emphasized factors that control
the translation of mRNA into protein, including the specific regulation of local
translation at synapses. Equally important have been studies of the influence of
reactive oxygen species (ROS) on the differentiation and maturation of neurons from
neural stem cells and of the basic factors that regulate gene transcription.
Vincent
Mauro and colleagues have been investigating the mechanisms by which eukaryotic
mRNAs initiate translation. Studies by Stephen Chappell and John Dresios revealed
that ribosomal recruitment can occur by Watson-Crick base pairing between complementary
segments of mRNA and ribosomal RNA and that such base pairing can facilitate the
nonlinear movement (shunting) of ribosomal subunits. Panagiotis Panopoulos is assessing
the physiologic relevance of such base-pairing interactions by blocking candidate
sites in the mRNA with complementary RNA oligonucleotides and assessing their effects
on the proteome.
Other studies from Dr. Mauro and his
colleagues have led to the development of new ideas about how ribosomal subunits
reach the initiation codon from the site of recruitment of the subunits. Dr. Chappell’s
studies with model mRNAs in transfected cells suggested that many instances of translation
initiation are more consistent with mechanisms involving tethering of ribosomal
complexes to the mRNA or with local ribosomal clustering than with the widely held
notion of ribosomal scanning. To test these hypotheses, Dr. Dresios is performing
in vitro studies to monitor the movement of ribosomal subunits along the mRNA 5′
leader. In addition, Dora Koh is developing a method to test a key prediction of
the tethering and clustering models, that is, that the accessibility of AUG codons
is an important factor affecting the use of the codons. She will monitor this accessibility
in living cells under various conditions that favor use of different AUG codons
in the 5′ leader of the mRNA encoding the β-secretase mRNA.
Bruce Cunningham and Peter Vanderklish
are defining the structure and function of a neuronal member of a unique family
of proteins that are upregulated during cold shock and other forms of stress. Working
with Annette Atkins and Julie Pillote, they have found that RNA-binding motif protein
3 (RBM3) is highly expressed in cerebellum, olfactory bulb, testis, and skin. The
protein appears in both the cytoplasm and the nucleus, and its distribution between
these compartments changes during development. In neurons, RBM3 is found in granules
that transport mRNA and components of the translation machinery into dendrites,
where the components can provide the basis for local translation of specific mRNAs
in response to synaptic activity. Much of the group’s efforts are currently
focused on characterizing the role of RBM3 in regulating local translation and on
determining the function of RBM3 in the nucleus, where it interacts with a variety
of other proteins and appears to be localized in novel subnuclear structures.
Appropriate regulation of gene expression
at the level of transcription is essential for normal mammalian development. Robyn
Meech and her colleagues are studying 2 transcription factors that play important
roles in the development of a diverse range of tissue types: the homeobox protein
Barx2 and the zinc finger neural restrictive silencer factor (NRSF).
Previous studies indicated that Barx2
regulates the differentiation of cartilage and muscle during embryonic development.
Recently, Katie Gonzalez found that mice that lack the gene for Barx2 (Barx2 null
mice) have impaired postnatal growth and have smaller muscles when they reach adulthood.
Moreover, she has found that Barx2 is expressed in satellite cells, suggesting that
it plays a role in muscle repair in adults. Dr. Meech and her colleagues have also
discovered a functional interaction between Barx2 and estrogen signaling. Moreover,
they found that Barx2 null mice have female-specific subfertility that may involve
a disruption of estrogen-regulated processes, including ovulation and embryo implantation.
This new role for Barx2 may lead to important insights into mechanisms of infertility.
NRSF is a transcription factor that represses
the expression of neuron-specific genes in nonneuronal cells. To understand this
mechanism better, Olivier Harismendy has developed a novel technology for genome-wide
analysis of transcription factor binding sites and has applied it to analysis of
NRSF targets during embryogenesis. Using this method, termed serial analysis of
genomic sites, he can identify transcription factor binding sites within the genomes
of intact cells or tissues. In conjunction with bioinformatic analysis, serial analysis
of genomic sites provides a high-throughput method to obtain information about the
essential DNA-protein interactions that regulate mammalian development.
Understanding the factors that support
neuronal differentiation and sustain neuronal survival are critical issues in neuronal
development. Kathryn Crossin and her colleagues previously showed that newborn neurons
have higher levels of ROS than their progenitors do, a surprising finding because
high levels of ROS are usually a reflection of dying cells. In more recent studies,
Marina Tsatmali showed that decreased ROS expression during the development of neurons
from multipotent progenitor cells dramatically altered neuronal morphology, biochemistry,
and physiology. She also found, using brain slices, that cells with high levels
of ROS were present at embryonic and early postnatal ages. The number of these cells,
however, decreased dramatically by postnatal day 20, except in regions such as the
hippocampus and olfactory bulb where neurogenesis takes place throughout life. The
cells with high levels of ROS had normal physiologic properties in situ comparable
to the physiologic properties of other neurons lacking high levels of ROS. Because
ROS are produced by mitochondria, we have hypothesized that alterations in mitochondrial
proteins and energetics are associated with the differentiation of different types
of neurons.
All of these efforts are designed to
elucidate the molecular and cellular events that define and regulate the development
and function of the nervous system. We are focusing on fundamental processes rather
than on specific diseases. This strategy is based on the notion that understanding
these processes can provide the necessary framework for defining the mechanisms
that underlie not just one but a variety of diseases. Indeed, our studies have already
provided insights into aspects of a number of diseases, including Alzheimer’s
disease, breast cancer, and mental retardation syndromes, as well as potentially
new bases for treatments related to the use of neural stem cells and for learning
and memory deficits.
Publications
Chappell, S.A., Dresios, J., Edelman, G.M., Mauro, V.P. Ribosomal
shunting mediated by a translational enhancer element that base pairs to 18S rRNA.
Proc. Natl. Acad. Sci. U. S. A. 103:9488, 2006.
Dresios, J., Chappell, S.A., Zhou, W., Mauro, V.P. An mRNA-rRNA
base-pairing mechanism for translation initiation in eukaryotes. Nat. Struct. Mol.
Biol. 13:30, 2006.
Stevens, T., Meech, R. BARX2 and estrogen receptor-α
(ESR1) coordinately regulate the production of alternatively spliced ESR1 isoforms
and control breast cancer cell growth and invasion. Oncogene 25:5426, 2006.
Tsatmali, M., Walcott, E.C., Makarenkova, H., Crossin, K.L. Reactive
oxygen species modulate the differentiation of neurons in clonal cortical cultures.
Mol. Cell. Neurosci. 33:345, 2006.
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