Small RNP Mediators of Gene Expression

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DESCRIPTION (provided by applicant): The nuclear steps of messenger RNA biogenesis are orchestrated by small nuclear ribonucleoproteins (snRNPs), which are central players in the action of two spliceosomes and the histone mRNA 3'-end processing machinery. Other classes of snRNPs guide the nucleotide modification of ribosomal RNAs in the nucleolus (snoRNPs) or of spliceosomal snRNAs in Cajal bodies (scaRNPs). A novel class of tiny RNPs, microRNPs, undergoes early biogenesis steps in the nucleus but functions in the cytoplasm to regulate the translation of mRNA targets. The mechanisms of action and biogenesis of snRNPs, scaRNPs and microRNPs will be investigated with an emphasis on the underlying molecular interactions using mammalian cells, the Xenopus oocyte, as well as Drosophila and yeast cells. How these small RNPs and their constituents contribute to the coupling of steps in gene expression is a central focus, including the synergies between and common components used to execute various processes. For box C/D scaRNPs, we will elucidate the signals and factors responsible for their Cajal body localization, which enables their action on spliceosomal snRNAs. We will investigate alternative splicing involving a choice between use of the major (U2-type) or minor (U12-type) spliceosome in the Drosophila prospero pre-mRNA. We will complete analyses of RNA functional groups and elucidate interactions with spliceosome components that are essential for the deposition of a multiprotein complex called the exon junction complex (EJC) on newly spliced mRNA upstream of exon-exon junctions. We will investigate alternative roles of a core component of the EJC (elF4AIII), which we have discovered can functionally replace its yeast homolog, Fall p. We will identify the subunit of the cleavage/polyadenylation factor CPSF that acts in 3'-end cleavage of histone pre- mRNAs, how the U7 snRNP recruits a subset of cleavage/polyadenylation factors, and how a conserved RNA element that binds U2 snRNP components stimulates histone mRNA 3'-end maturation. We will investigate how microRNP components such as AGO2 and FXR1 play repressive versus stimulatory roles in translation depending on cell growth conditions, and whether the processing of microRNA primary transcripts occurs in discrete nucleoplasmic loci. These studies will illuminate a variety of human disease processes that can be traced to perturbations in small RNP biogenesis, functions and associations.