The goal of this project is to understand the coordination of histone mRNA stability with DNA replication during the m ammalian cell cycle. The experiments included in this proposal are designed to: a) identify and delineate the specific regions of cell cycle-regulated histone messenger RNAs responsible for coupling metabolic stability with DNA synthesis, and b) characterize the molecules and mechanisms involved in the rapid degradation of histone mRNA that occurs in the absence of DNA replication. Fusion genes will be constructed containing cell cycle regulated histone genes under control of a human heat shock gene promoter to allow transcriptional and post-transcriptional regulation to be dissociated. The effects of specific modifications introduced into the histone gene sequence on stability of transcripts during the cell cycle and following inhibition of DNA synthesis will be determined in order to identify sequences involved in regulating mRNA turnover. In addition, equivalent regions of a cell cycle dependent and a cell cycle independent human H4 histone gene will be exchanged. Analysis of the behavior of the resulting chimeric transcripts will indicate the contribution of 5' and 3' domains toward mRNA stability. To gain insight into the mechanism responsible for selective histone mRNA turnover, experiments will be conducted to determine whether the mechanism: 1) operates throughout the cell cycle, 2) is localized to a specific cytoplasmic compartment, 3) is specific for each class of histone mRNA, and 4) involves changes in histone mRNA-protein interactions. Finally, an in vitro system will be developed to allow the assay, identification and characterization of factors mediating histone mRNA turnover. These studies should lead to a better understanding of how histone gene expression is modulated during the cell cycle. Moreover, the results will be relevant to the broader problem of control of mRNA turnover, which is now recognized as an important level of gene regulation during development and in proliferative diseases.
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