The steady-state level of specific mRNA is determined by the rates by which this molecule is synthesized and degraded. Despite the importance of mRNA turn-over in the control of gene expression, the mechanisms that underlie this process are still obscure. It is thus the long-term goal of the proposed research to explore the mechanism of intracellular mRNA breakdown. In recent studies the PI has shown that the breakdown rate of the mRNA for PEP- carboxykinase (PEPCK) is hormonally regulated and that cAMP (as an intracellular mediator of glucagon) stabilizes the mRNA against degradation. It has also been demonstrated that 733 nucleotides from the 3'-region of the mRNA are sufficient to confer the cAMP regulation of mRNA turn-over. The main goal of this proposal is to delineate the nucleotide domains that mediate the cAMP regulation of PEPCK mRNA as well as the determinants that confer the short half-life of PEPCK mRNA. These studies will involve monitoring the half-lives of mRNA transcripts of a variety of hybrid genes containing different sequences of PEPCK, after these genes have been introduced into hepatoma cells. In addition, we propose to examine the role of RNA-protein interaction at the cAMP- responsive element in the control of the half-life of PEPCK mRNA. It is hoped that these studies will result in a better understanding of the molecular events which control gene expression and also will find a practical use in the construction of an efficient expression system in which the control of mRNA stability is considered when a foreign gene is to be introduced into cells.//
