Poster abstracts
Poster number 92 submitted by Katherine Sortino
RESC8 and RESC14 cooperate to mediate RESC function and dynamics during trypanosome RNA editing
Katherine Sortino (Department of Microbiology & Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, Buffalo, NY 14203.), Brianna Tylec (Department of Microbiology & Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, Buffalo, NY 14203.), Natalie M. McAdams (Department of Microbiology & Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, Buffalo, NY 14203.), Runpu Chen (Department of Microbiology & Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, Buffalo, NY 14203.), Yijun Sun (Department of Microbiology & Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, Buffalo, NY 14203.), Laurie K. Read (Department of Microbiology & Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, Buffalo, NY 14203.)
Abstract:
Uridine insertion/deletion RNA editing is unique to kinetoplastid organisms and essential for their survival and virulence. This intricate process requires the action of several multiprotein complexes and small trans-acting RNAs called guide RNAs. The RNA editing holoenzyme comprises three dynamically interacting complexes: RECC (RNA Editing Catalytic Complex), RESC (RNA Editing Substrate Binding Complex), and REH2C (RNA Editing Helicase 2 Complex). Previous studies in Trypanosoma brucei showed that RESC components, RESC14 and RESC8, are critical in maintaining normal protein-protein and protein-RNA interactions within the editing holoenzyme. To further clarify the role of these proteins and the RESC complex in RNA editing, we performed high-throughput sequencing of ATP synthase subunit 6 (A6) mRNA in RESC14, RESC8, and RESC13 RNAi cells, and compared the phenotypes of these knockdown cell lines. We measured a highly significant overlap between exacerbated pause sites arising on A6 mRNA in cells depleted of RESC14 and RESC8, suggesting an overlapping function between these proteins during editing progression. In contrast, A6 mRNA from RESC13 depleted cells exhibited different pause sites, indicative of a function distinct from that of RESC14/8. Additional features of editing progression, such as the lengths of mis-edited junctions, also differed between RESC14/8 and RESC13, supporting their specific functions and suggesting that both RESC14 and RESC8 are needed for junction formation on A6 mRNA. To probe the biochemical basis of the RESC14/8 interdependence, we performed Native PAGE analysis and showed that RESC8 is not incorporated into large editing complexes in the absence of RESC14. Co-immunoprecipitation assays also identified an RNA inhibited interaction between the RESC14 and RESC8. Together, our findings suggest that RESC14 and RESC8 rely on each other for necessary protein and RNA rearrangements during editing progression. Our working model is presented.
Keywords: Trypanosome, RNA editing