The goal of this R03 research proposal is to expand our understanding of the basic molecular mechanism(s) regulating expression of the mouse somatostatin receptor subtype 5 gene (sst5).
The aim i s to identify and clone cellular factors that interact with the sst5 promoter and characterize their functional importance in pituitary cells. Specific emphasis will be placed on the cellular factors important for sst5 promoter activity in TtT97 thyrotropes. These experiments will be a direct extension of the work previously funded by a K08 Mentored Clinical Scientist Development Award. The overall goal of this work is to gain a better understanding of the interaction between the hypothalamicpituitary-thyroid axis and the somatostatinergic system and their combined effect on pituitary thyrotrope regulation. Using the murine TtT-97 thyrotropic tumor model, we have previously demonstrated that treatment with thyroid hormone reduces tumor size in association with enhanced expression of the sst5 gene. Studies performed as part of the K08 Award have initially characterized these in vivo observations by investigating basal promoter activity of the sst5 gene and its regulation by thyroid hormone. The mouse sst5 gene has been cloned, its transcription start site localized and its structure identified as having 3 exons and 2 introns. Functional promoter activity has been assessed in transient transfection experiments in a variety of pituitary cell lines and primary TtT97 thryotrope cultures. Sequences important for basal promoter activity have been identified using a deletion and mutational strategy. The region between -67 and -47 appear to contain a nucleotide sequence that is critical for basal trancriptional regulation of the sst5 gene in pituitary cells. The functional importance of this region has been confirmed by DNase I protection analysis to identify DNA protein interactions. Sequence analysis of these critical sst5 promoter regions revealed nucleotide homology to potential binding sites for known transcription factors, including Ptx 1 and Egr-1. The role of these factors and others in regulation of the mouse sst5 promoter will be the subject of this R03 proposal. Not only will these data enhance our understanding of the basic molecular mechanisms of the somatostatinergic system and pituitary regulation, but have long range clinical applications regarding the pathogenesis and treatment of neuroendocrine tumors. The somatostatin analog, octreotide, has been used to treat a variety of neuroendocrine tumors and description of the molecular mechanisms of the somatostatinergic system will be of clinical application to these diseases. Further knowledge of somatostatin receptor function and regulation will potentially contribute to the development of more efficacious somatostatin analogs for the diagnosis and treatment of neuroendocrine tumors. Additionally, understanding the molecular mechanism(s) underlying transcriptional regulation of these receptors may contribute to our ability to manipulate somatostatin receptor status in specific tumors and enhance clinical response to somatostatin analog therapy.
