This project will reveal the distinct way in which proteins are made in the Bacteroidetes, a family of bacteria that are found in the healthy human intestine as well as in many natural habitats. Bacteroidetes lacks the Shine-Dalgarno sequence, a well-known control elelment that is required for translation of RNA into proteins in most other bacteria, and this research is aimed at determining what takes its place in this bacterial lineage. This project will also provide (1) research training for undergraduate, graduate, and postdoctoral students, (2) summer research opportunities for local high school students (Columbus Public Schools), and (3) faculty mentorship for students of the Louis Stokes Alliance for Minority Participation (LSAMP) program, which aims to boost the success rate of underrepresented minority students in STEM (Science, Technology, Engineering, and Mathematics) disciplines.
Initiation of translation involves assembly of a ribosome complex in which the initiator tRNA is paired to the start codon of mRNA. The fidelity of translation depends on recognition of the correct start codon among all AUG and similar trinucleotide sequences in the mRNA. In E. coli and many bacteria, recognition of the start codon is facilitated in part by base pairing between the Shine-Dalgarno (SD) sequence of the mRNA (which lies just upstream from the start codon), and the anti-Shine-Dalgarno (ASD) sequence of the 30S subunit (contained near the 3? end of the 16S rRNA). Recent genome sequencing studies have revealed that whole taxonomic groups of bacteria completely lack SD sequences, despite that the ASD sequence remains intact and conserved in these organisms. These include the phyla Bacteroidetes and a subset of Cyanobacteria. The primary goals of this project are to uncover the sequence determinants of the translation initiation regions (TIRs) in the Bacteroidetes and understand how these TIR sequences govern the rate of initiation. Modern techniques such as ribosome profiling will be used in conjunction with biochemical approaches to achieve these goals. Features of the TIR aside from the SD, such as secondary structure and S1 binding sites, are also thought to impact initiation rate in bacteria. By studying the Bacteriodetes system, the roles of these features may be clarified, as the complication of SD-ASD pairing is effectively removed.
