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The Skaggs Institute
for Chemical Biology


Scientific Report 2006




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.

 

Gerald M. Edelman, M.D., Ph.D.
Professor
Chairman, Department of Neurobiology