Poster abstracts
Poster number 99 submitted by Fawwaz Naeem
Analysis of programmed frameshifting during translation of prfB in Flavobacterium johnsoniae
Fawwaz M. Naeem (Ohio State Biochemistry Program, Center for RNA Biology, OSU), Bryan T. Gemler (Interdisciplinary Biophysics Graduate Program, Center for RNA Biology, OSU), Zakkary A. McNutt (Ohio State Biochemistry Program, Center for RNA Biology, OSU), Ralf Bundschuh (Interdisciplinary Biophysics Graduate Program, Center for RNA Biology, Department of Physics, Department of Chemistry & Biochemistry, Division of Hematology, Department of Internal Medicine, OSU), Kurt Fredrick (Ohio State Biochemistry Program, Center for RNA Biology, Microbiology Department, OSU)
Abstract:
Ribosomes of Bacteroidia fail to recognize Shine-Dalgarno (SD) sequences due to sequestration of the 3’ tail of the 16S rRNA on the 30S platform. Yet in these organisms, the prfB gene typically contains the programmed +1 frameshift site with its characteristic SD sequence. Here we investigate prfB autoregulation in Flavobacterium johnsoniae, a member of the Bacteroidia. We find that the efficiency of prfB frameshifting in F. johnsoniae is low (~7%) relative to that in E. coli (~50%). Mutation or truncation of bS21 in F. johnsoniae increases frameshifting substantially, suggesting that anti-SD (ASD) sequestration is responsible for the reduced efficiency. The frameshift site of certain Flavobacteriales, such as Winogradskyella psychrotolerans, lacks the SD sequence. In F. johnsoniae, this W. psychrotolerans sequence supports frameshifting as well as the native sequence, and mutation of bS21 causes no enhancement. These data suggest that prfB frameshifting normally occurs without SD-ASD pairing, at least under optimal laboratory growth conditions. Chromosomal mutations that ablate the SD and remove the frameshift confer subtle growth defects in the presence of streptomycin or paraquat, respectively, indicating that both the autoregulatory mechanism and the SD element contribute to F. johnsoniae cell fitness. Analysis of prfB frameshift site across 2686 representative bacteria shows loss of the SD sequence in many clades, with no obvious relationship to genome-wide SD usage. These data reveal unexpected variation in the mechanism of frameshifting and identify another group of organisms, the Verrucomicrobiales, that globally lack SD sequences.
References:
Baranov PV, Gesteland RF, Atkins JF. 2002. Release factor 2 frameshifting sites in different bacteria. EMBO Rep 3: 373–377
Craigen WJ, Caskey CT. 1986. Expression of peptide chain release factor 2 requires high-efficiency frameshift. Nature 322: 273–275
Jha V, Roy B, Jahagirdar D, McNutt ZA, Shatoff EA, Boleratz BL, Watkins DE, Bundschuh R, Basu K, Ortega J, et al. 2021. Structural basis of sequestration of the anti-Shine-Dalgarno sequence in the Bacteroidetes ribosome. Nucleic Acids Res 49: 547–567
McNutt ZA, Roy B, Gemler BT, Shatoff EA, Moon K-M, Foster LJ, Bundschuh R, Fredrick K. 2023. Ribosomes lacking bS21 gain function to regulate protein synthesis in Flavobacterium johnsoniae. Nucleic Acids Res 51: 1927–1942
Keywords: prfB +1 programmed frameshifting, Flavobacterium johnsoniae , Ribosomes