Coupling of transcription elongation and termination with pre-mRNA processing

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Project Summary/Abstract The production of messenger RNA (mRNA) is the primary event in gene expression where genetic information is transfered from the gene’s DNA into a disposable RNA copy. Corruption of this process is a hallmark of many diseases including cancer. mRNA synthesis requires not only synthesis of an RNA transcript but maturation of that transcript by 5’ capping, excision of introns and splicing of exons and 3’ end formation by cleavage and polyA tail addition. The mRNA processing steps that radically transform the primary transcript occur largely co-transcriptionally; that is to say the substrate of mRNA processing is the growing nascent RNA that is extruded by an RNA polymerase II (pol II) molecule at rates of 500-5000 bases/min. Our working model is that synthesis and processing of a mRNA precursor are carried out in an integrated fashion within a dynamic 'mRNA factory' complex that includes both RNA polymerase and processing factors some of which make direct contacts with the pol II C-terminal domain (CTD). The goal of our work is to understand how growth of the RNA chain by transcription is coordinated with its folding into RNA secondary structures, its association with RNA binding proteins, and its maturation by splicing and 3’ end formation. These important features of nascent RNA metabolism are all affected by how fast the RNA chain grows. Therefore it is important to discover how the speed of pol II is controlled as it travels along genes. When pol II completes its journey to the end of the gene, the highly stable transcription elongation complex must be actively disassembled to recycle the enzyme and prevent it from invading neighboring genes. We will investigate how the process of transcription termination is achieved in carefully controlled ways at the 3’ ends of genes and also within genes where termination can occur “prematurely”. We will use genetic and genomic approaches in human cells to investigate these three Key Challenges: I. What is the relation between pre-mRNA processing, nascent RNA folding, RNA binding protein (RBP) binding, and transcription elongation? II. How is the speed of transcription elongation regulated? III. What mechanisms terminate pol II transcription within genes and downstream of genes?