This project proposes to study the mechanism by which cell-specific expression of the neuro-and endocrine peptides calcitonin and calcitonin gene-related peptide (CGRP) is determined at a transcriptional and post- transcriptional level. Cell-specific alternative RNA processing events direct the preferential expression of CGRP mRNA from the rat calcitonin gen in neurons of the peripheral and central nervous system, where the encoded peptide functions in sensory, motor and integrative systems. Four areas of research are proposed: 1) Regulation of alternative splicing using mutagenesis and gene transfer studies. 2) Somatic producing cells will be analyzed to test the hypothesis that a neuron-specific factor is required t produce CGRP mRNA. 3) Analysis of cell-free extracts in vitro. The ability of factors in nuclear extracts from tissue culture cell lines to direct alternative splicing of the calcitonin gene will be studied by using in vit o splicing assays, in vitro RNA-factor binding gel retardation assays. In addition extracts will be analyzed for the presence of cell-specific differences in the presence of small nuclear RNAs (snRNAs) and their associated polypeptides (snRNP proteins). 4) Identification of cell specifi regulatory elements/enhancers for the calcitonin/CGRP gene. Sequences associated with the calcitonin gene and its flanking DNA will be studied to identify elements that promote enhanced transcription in medullary thyroid carcinoma cell (MTC) and ultimately neuronal cells. This information will b used to conduct comparative analyses between enhancer elements and factors of the calcitonin/CGRP gene and other neuropeptide genes that are coexpress d in MTCs and some peripheral neurons such as preprotachyleinin somatostatin and cholecystokinin. The award period will be used to further develop the candidate as an independent researcher within the environment of a highly interactive Pharmacology department and School of Medicine. This environme t is ideal for the study of neuroendocrine gene expression as it has strong research faculty in cell and molecular biology, biochemistry and neurobiology. The RCDA, if awarded, would allow the candidate to commit on y a modest amount of time to teaching in medical school courses, and it would promote the candidates development as an independent research scientist in the Medical School. In the long-term, the program of study proposed could allow the candidate to pursue the development of novel pharmacological tool ; such as nucleic acids based drugs, that are designed to regulate the abnorm l expression of specific genes that may arise in pathological states.
| Roesser, J R; Liittschwager, K; Leff, S E (1993) Regulation of tissue-specific splicing of the calcitonin/calcitonin gene-related peptide gene by RNA-binding proteins. J Biol Chem 268:8366-75 |
