Defining the molecular mechanisms by which specific genes in the neuroendocrine system are activated, and how their expression is regulated in terminally differentiated cells, is a central problem in the field of endocrinology. A major goal under this grant has been to identify cell- specific strategies by which neuroendocrine genes are regulated, particularly in hypothalamus and in the diverse neural crest- derived endocrine and non-endocrine cell types in which genes encoding regulatory peptides are expressed. In the previous project period, the principal investigator discovered and characterized several novel POU domain transcription factors expressed in specific hypothalamic cell types characterized by expression of corticotropin-releasing hormone (CRH) and neural-crest derived cell types expressing calcitonin gene related peptides (CGRP). These transcription factors could serve as determining factors in hypothalamus and neural crest development. In this competitive renewal, focus is on the investigation of the developmental roles of these putative determining factors in activation of specific genes of the neuroendocrine system, using homologous recombination in embryonic stem cells to generate mice null for each transcription factor genomic locus. It is hypothesized that these POU domain factors serve critical functions both in activation of the CRH and CGRP transcription units, exerting roles during ontogeny, terminal differentiation of cell phenotypes, and cell viability. Their roles will be assessed in activation of other critical transcription units of the neuroendocrine system including vasopressin, oxytocin, GRH, and substance P. Based on initial data, these experiments are likely to genetically link the novel transcription factors to control of the hypothalamic- pituitary axis and regulatory genes of the neuroendocrine system. Experiments will be directed at investigating the basis for selective actions of highly related factors in target gene activation, and the molecular mechanisms by which these POU domain factors select highly restricted bipartite DNA binding sites. These experiments will provide insights into developmental strategies involved in the endocrine hypothalamus, in activating specific regulatory genes, and in generating mature cell types in the neuroendocrine system.
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