Structural and functional studies of mRNA processing and quality control

Abstract

Eukaryotic mRNA precursors (pre‐mRNAs) must undergo extensive maturational processing, including 5′‐end capping, splicing and 3′‐end cleavage and polyadenylation. Turnover of mature mRNAs is mediated by exoribonucleases that function in the 5′→3′ or 3′→5′ direction (as well as other enzymes), which are important for the degradation of defective mRNAs as well (mRNA quality control). A nuclear 5′→3′ exoribonuclease (XRN2, Rat1 in yeast) also couples pre‐mRNA 3′‐end processing and Pol II transcription termination.We have been studying the structure and function of various components of the 3′‐end processing machinery as well as the 5′→3′ exoribonucleases. For example, we showed that Ssu72, a Pol II C‐terminal domain (CTD) pSer5 phosphatase, binds a pSer5 peptide with the pSer5‐Pro6 peptide bond in the cis configuration. Moreover, Ssu72 also recognizes the CTD phosphorylated at Ser7, but a pSer7 peptide is bound in the opposite direction as compared to the pSer5 peptide.Unexpectedly, our studies on a protein factor (Rai1) that stimulates the 5′→3′ exoribonuclease Rat1 led to the discovery of a novel quality control mechanism for mRNA 5′‐end capping. Studies in yeast and mammalian cells confirm that mRNAs with incomplete 5′‐end caps are produced under physiological conditions, and that Rai1 and its mammalian homolog (DXO) have important roles in the detection and degradation of such defective mRNAs.