The rigid coupling of pancreatic insulin production to blood glucose concentration is the keystone of mammalian metabolic homeostasis with even minor alterations in this relationship jeopardizing survival. While the nature of the process by which glucose stimulates insulin secretion and biosynthesis is poorly understood, control is known to be exerted at both the steps of protein translation and mRNA transcription. Because individual genes contain discrete sequence that encode their own transcriptional control program and modern techniques have allowed for the identification of these regulatory elements, the insulin gene might likely contain distinct targets of the glucose response pathway(s). In transfection experiments using cultured hamster insulinoma (HIT) cells, glucose stimulates the islet-cell specific rat insulin-I promoter driven expression of unrelated reporter genes. This effect has been recently confirmed in primary fetal rat islet cell cultures. Based upon these findings, this proposal will seek to pinpoint """"""""glucose response elements"""""""" by studying the activity of mutated insulin promoters in HIT and fetal islet cells. These putative regulatory elements will be evaluated for the capacity to bind islet cell factors that may serve as the mediators of metabolic control. Preliminary studies have revealed that in HIT cells, a short 50 base pair segment harbors both islet-specificity and glucose control as well as being a domain of complex protein-binding activity. Utilizing the glucose response element(s) as binding probes, the ultimate goal of this proposal is to clone these regulatory factors from islet cell cDNA libraries. It is hoped that, once characterized, these DNA binding proteins will constitute the most distal components of a glucose/metabolic pathway that might converge to a common mechanism also responsible for insulin secretory and translational control. The cloned factors may become useful tools with which to dissect this system.
| Amsterdam, A; Lin, S; Moss, L G et al. (1996) Requirements for green fluorescent protein detection in transgenic zebrafish embryos. Gene 173:99-103 |
| Peyton, M; Moss, L G; Tsai, M J (1994) Two distinct class A helix-loop-helix transcription factors, E2A and BETA1, form separate DNA binding complexes on the insulin gene E box. J Biol Chem 269:25936-41 |
| Emens, L A; Landers, D W; Moss, L G (1992) Hepatocyte nuclear factor 1 alpha is expressed in a hamster insulinoma line and transactivates the rat insulin I gene. Proc Natl Acad Sci U S A 89:7300-4 |
| German, M S; Moss, L G; Wang, J et al. (1992) The insulin and islet amyloid polypeptide genes contain similar cell-specific promoter elements that bind identical beta-cell nuclear complexes. Mol Cell Biol 12:1777-88 |
