Studies of translation initiation in bacteria

这个项目将揭示拟杆菌门中蛋白质形成的独特方式，拟杆菌门是一种在健康人类肠道和许多自然栖息地中发现的细菌家族。拟杆菌门缺乏Shine-Dalgarno序列，这是一种在大多数其他细菌中将RNA翻译成蛋白质所需的众所周知的控制元件，这项研究旨在确定在这种细菌谱系中占据什么位置。该项目还将提供(1)为本科生、研究生和博士后提供研究培训，(2)为当地高中生（哥伦布公立学校）提供暑期研究机会，以及(3)为路易斯·斯托克斯少数民族参与联盟（LSAMP）项目的学生提供教师指导，该项目旨在提高少数民族学生在STEM（科学、技术、工程和数学）学科中的成功率。翻译的起始涉及核糖体复合物的组装，其中启动tRNA与mRNA的起始密码子配对。翻译的保真度取决于对所有AUG和mRNA中相似的三核苷酸序列的正确起始密码子的识别。在大肠杆菌和许多细菌中，起始密码子的识别部分是通过mRNA的Shine-Dalgarno （SD）序列（位于起始密码子的上游）和30S亚基的anti-Shine-Dalgarno （ASD）序列(位于3？16S rRNA末端)。最近的基因组测序研究表明，尽管ASD序列在这些生物中保持完整和保守，但整个细菌分类群完全缺乏SD序列。这些包括拟杆菌门和蓝藻的一个子集。该项目的主要目标是揭示拟杆菌门翻译起始区（TIR）的序列决定因素，并了解这些TIR序列如何控制起始率。核糖体分析等现代技术将与生化方法结合使用，以实现这些目标。除SD外，TIR的特征，如二级结构和S1结合位点，也被认为会影响细菌的起始率。通过对Bacteriodetes系统的研究，可以明确这些特征的作用，有效地消除SD-ASD配对的复杂性。

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.