This project includes several studies on the genetic regulation of the biosynthesis of protein precursors of neuropeptides in the mammalian nervous system. A. In cholestatic liver disease plasma enkephalin levels are elevated, and the cholestasis-associated pruritus (itching) is ameliorated by opiate antagonists, suggesting that endogenous opioid peptide biosynthesis, degradation, or release may be altered in cholestasis. To test this hypothesis, we measured levels of mRNAs coding for the three opioid peptide precursors in the adult rat and human liver, an organ which normally produces no opioid peptides. We found that experimental cholestasis in rat elicited by bile duct resection strongly induces proenkephalin (proEnk) mRNA, but not prodynorphin or proopiomelanocortin mRNAs, in liver. In situ hybridization revealed that proliferating bile ductular cells, rather than hepatocytes, are the locus of proEnk mRNA. In spite of this finding in rat, proEnk mRNA was not detected in liver biopsy samples of several patients with cholestatic liver disease; however, a role for proEnk induction in the pathology of cholestatic disease is still under investigation. B. The transactivator protein tax1 of the human T-cell leukemia virus I (HTLV-I) was found to transactivate the proEnk gene in a glial cell line. Sequences between bases -437 and -190 relative to the transcription start site were necessary for maximal transactivation. This finding may be relevant to HTLV-I-caused diseases, such as T-cell leukemia and tropical spastic paraparesis, in which proEnk-producing cells are infected. C. Ongoing studies on the regulation of the gene coding for the precursor of neuropeptide Y (NPY), an important peptide neurotransmitter in both central and peripheral nervous systems, revealed that a cell-specific cyclic AMP/phorbol ester inducible element may exist in the 5'flanking sequence of the rat NPY gene between bases -2300 and -1800. However, sequences outside the region of the promoter tested (-2300 to +17) appear to be responsible for neuron-specific expression of this gene. These studies will hopefully shed light on the control of biosynthesis of peptides that are important in autonomic regulation, pain perception, and cognitive function.
