This project continues an on-going, well developed, diverse research program applying biochemistry, molecular, cellular and structural biology methodologies to understand the biological implications of a nuclear RNA decapping protein. The aims of the project are (i) to identify the molecular interactions between RNA substrates and the nuclear decapping protein and perform structure-based mutagenesis to validate these and (ii) to identify authentic in vivo RNA substrates for this protein and demonstrate that alterations in the amount of protein correlates with changes in target RNA stability in vivo. Collectively, these will provide an understanding of the mechanism of substrate recognition and catalytic activity of the X29 decapping enzyme, a protein that is evolutionarily well conserved and has functional homologues in all vertebrates. Previous work demonstrated that the frog and human proteins hydrolyze 5' monomethylated caps from RNAs releasing m7GDP and leaving a 5' monophosphate on the RNA. In vivo such decapped RNAs would be degraded by 5'-3' exonucleases. The hypothesis of this project is that the X29 protein regulates the levels of nuclear RNAs in vivo and may specifically degrade subclasses of RNAs that may not be correctly spliced, modified or transported. This project will determine whether X29 is a previously unknown nuclear counterpart of the cytoplasmic decapping complex and directs 5'-3' decay of nuclear RNAs altering the in vivo levels of mRNAs as well as RNAs involved in ribosome biogenesis, pre-mRNA splicing and mRNA export.
A significant amount of the research will be performed by graduate students and undergraduates at MU who will work directly with the PI in performing experiments and interpreting data. This one-on-one training/education environment is empowering to young scientists; group lab meetings and informal discussions facilitate and reinforce peer-learning. Their own work allows them to see and understand how various 'specialized' areas of science are complementary but together can provide unique insight into otherwise intractable questions. The pool of undergraduate students from which this laboratory currently employs include MU Biochemistry majors looking for a research experience for their honors thesis, summer LSUROP (Life Sciences Undergraduate Research Program) recruits (minorities and undergrads from small liberal arts colleges across the US living at MU for 8 weeks over the summer) and the EXPRESS program at MU which facilitates minority students enrolled at MU in obtaining experience in a research lab.
