Coupling of Transcription with Pre-mRNA Metabolism

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): The synthesis of messenger RNA is the primary event in gene expression and is central to the life of cells. mRNA production requires transcription by pol II and processing of the primary transcript by a set of proteins which carry out capping, splicing and cleavage/polyadenylation. Transcripts made in by polymerases other than pol II are not processed correctly into mature mRNA. A network of protein:protein contacts in the cell nucleus exists to couple transcription by pol II with the specific RNA processing events responsible for maturation of mRNA. The objective of this proposal is to elucidate how proteins communicate to achieve coupling of pol II transcription with RNA packaging and processing using biochemical and genetic approaches in mammalian cells, frog oocytes and budding yeast. Our research is testing the idea that mRNA is made by a "factory" complex containing RNA pol II and RNA processing factors which contact it through a repetitive protein domain called the CTD. This model has changed the way we think about proteins which were once thought to operate independently but are now thought to be co-ordinated with one another in the nucleus. One important functional consequence of the integration between transcription and processing that we are beginning to uncover is that transcription factors can regulate the efficiency of RNA processing and conversely processing factors can potentially regulate transcription. This work may help elucidate how the transcription and processing of mRNAs are regulated under normal conditions and how they become mis-regulated in the disease state. Defects in splicing resulting of pre-mRNA's are responsible for a large fraction of all inherited diseases. The specific aims of this work are:1. To compare mechanisms of RNA processing and packaging that are coupled to pol II transcription with uncoupled processing and packaging of RNA in vivo.2. To determine how how capping enzymes regulate pol II transcription and how components of the pol II transcription machinery regulate capping.3. To determine how pol II elongation rate, CTD heptad sequences and the CTD C-terminus affect RNA packaging, capping, splicing and 3' end formation.4. To determine whether mammalian processing factors associate with elongating pol II ternary complexes in vivo and how the processing factor composition of ternary complexes changes during elongation?