Pheochromocytoma cells in vitro and in vivo may exhibit some phenotypic characteristics which are """"""""appropriate"""""""" functions of normal chromaffin cells and others that are not expressed by normal chromaffin cells. The latter are generally highly differentiated characteristics of a variety of amine and peptide hormone-producing neural and endocrine cells from other parts of the body, rather than expressions of random genomic derepression. The molecular mechanisms that cause pheochromocytoma cells to exhibit aberrant characteristics selected from a finite repertoire of neuroendocrine functions and the events that lead to expression of these characteristics in the course of neoplastic progression are not known. Our studies at the cellular level indicate that production of a number of appropriate and ectopic neuropeptides is regulated by corticosteroids and/or NGF in PC12 rat pheochromocytoma cells or in human pheochromocytomas. Some of these peptides are also produced by normal chromaffin cells in vitro but not in vivo. We will now extend our studies to the molecular level to investigate mechanisms involved in regulation of these markers. Using mRNA from PC12 cells as templates, we will construct and clone complementary DNAs (cDNAs) encoding a number of neuropeptides. These cloning efforts will begin with neurotensin (NT), a functional marker applicable to both human and rat pheochromocytomas. These cDNAs will be used in hybridization assays to quantitate changes in specific mRNA levels in response to corticosteroids and NGF and to investigate the organization of the neuropeptide genes. They will also permit us to determine the complete amino acid sequences of the neuropeptide precursors. We will concomitantly expand our studies at the cellular level, comparing mechanisms that regulate the synthesis, storage, and secretion of NT and other neuropeptides in normal, hyperplastic, and neoplastic chromaffin cells of humans and rats. These studies will shed light on mechanisms that determine normal neuroendocrine cell phenotype and on ways in which normal regulation might become deranged during the development of neoplasia. (B)
