Our research interest is to understand the molecular mechanism of the pre-mRNA splicing reaction, using the budding yeast Saccharomyces cerevisiae as a model system. Splicing is a fundamental process of eukaryotic gene expression. Many diseases are associated with splicing defects or mis-regulation of splicing. The splicing reaction takes place on the spliceosome assembled via ordered interactions with the pre-mRNA of five snRNAs and numerous protein factors. The snRNAs play roles in the recognition and alignment of the splice sites through base pairing with the intron sequence and with each other to form the catalytic core. The protein factors function to facilitate and stabilize RNA base pairing, and also mediate interactions between splic sites and structural changes of the spliceosome between assembly steps.
Our research focuses on understanding the functional roles of protein splicing factors through identification and characterization of these factors using biochemical methods. Throughout the years, we have identified and uncovered the functions of several protein factors and protein complexes involved in various steps of the pathway. These include the NTC required for spliceosome activation, the NTR complex for spliceosome disassembly, and four proteins, Yju2, Cwc22, Cwc24 and Cwc25, essential for the first catalytic reaction. We have generated many tools for further dissection of the splicing pathway through characterization of these components, and are now able to arrest splicing at different stages of the pathway and purify different spliceosome intermediates for mechanistic studies of the reaction. Our work has revealed dynamic properties of the spliceosome for its catalytic capacity and for interactions between the proteins and the splice site at different steps. We have also elucidated the mechanisms of how DExD/H-box RNA helicases may mediate remodeling of the spliceosome in facilitating the splicing reaction and in splicing fidelity control. Our studies have provided many new mechanistic insights into the mechanism of the splicing reaction.
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