The replication dependent histone mRNAs are the only metazoan mRNAs that are not polyadenylated. Instead they end in a conserved stemloop structure. The synthesis of histone proteins is tightly coupled to DNA replication and this is controlled largely by regulating histone mRNA. In early embryos, all organisms must produce histone proteins in the absence of transcription, and many embryos, including frogs and Drosophila, must produce massive amounts of histone proteins in a short time during the extremely rapid early cell divisions. We have carried out a genome wide RNAi screen for factors involved in histone mRNA processing using Drosophila cultured cells. We will determine the function of these factors in the processing reaction (components of the U7 snRNP and the cleavage factor) as well as other possible previously unknown factors, and identify which factors are present in the Histone Locus Body. We also obtained factors involved in replication-independent chromatin assembly in the screen suggesting that the pathways for replication-dependent and replication-independent histone protein metabolism directly interact. We will determine the role of these factors in histone mRNA regulation in the fly, using available mutants in many of these factors. We will also determine the molecular mechanism of regulation of histone mRNA translation in Xenopus oocytes, focusing on the role of the stemloop binding protein, and a novel protein that it interacts with, in translation. This novel protein, SLIP1, is likely involved in translation regulation of other mRNAs, and we will identify what other pathways it participates in.
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