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mBio

Title:		Discovery of Unannotated Small Open Reading Frames in Streptococcus pneumoniae D39 Involved in Quorum Sensing and Virulence Using Ribosome Profiling
Author(s):		Laczkovich I; Mangano K; Shao X; Hockenberry AJ; Gao Y; Mankin A; Vazquez-Laslop N; Federle MJ;
Address:		"Department of Microbiology and Immunology, University of Illinois at Chicagogrid.185648.6, Chicago, Illinois, USA. Center for Biomolecular Sciences, University of Illinois at Chicagogrid.185648.6, Chicago, Illinois, USA. Department of Pharmaceutical Sciences, University of Illinois at Chicagogrid.185648.6, Chicago, Illinois, USA. Department of Integrative Biology, University of Texas at Austin, Austin, Texas, USA"
Journal Title:		mBio
Year:		2022
Volume:		20220719
Issue:		4
Page Number:		e0124722 -
DOI:		10.1128/mbio.01247-22
ISSN/ISBN:		2150-7511 (Electronic)
Abstract:		"Streptococcus pneumoniae, an opportunistic human pathogen, is the leading cause of community-acquired pneumonia and an agent of otitis media, septicemia, and meningitis. Although genomic and transcriptomic studies of S. pneumoniae have provided detailed perspectives on gene content and expression programs, they have lacked information pertaining to the translational landscape, particularly at a resolution that identifies commonly overlooked small open reading frames (sORFs), whose importance is increasingly realized in metabolism, regulation, and virulence. To identify protein-coding sORFs in S. pneumoniae, antibiotic-enhanced ribosome profiling was conducted. Using translation inhibitors, 114 novel sORFs were detected, and the expression of a subset of them was experimentally validated. Two loci associated with virulence and quorum sensing were examined in deeper detail. One such sORF, rio3, overlaps with the noncoding RNA srf-02 that was previously implicated in pathogenesis. Targeted mutagenesis parsing rio3 from srf-02 revealed that rio3 is responsible for the fitness defect seen in a murine nasopharyngeal colonization model. Additionally, two novel sORFs located adjacent to the quorum sensing receptor rgg1518 were found to impact regulatory activity. Our findings emphasize the importance of sORFs present in the genomes of pathogenic bacteria and underscore the utility of ribosome profiling for identifying the bacterial translatome. IMPORTANCE This work employed pleuromutilin-assisted ribosome profiling using retapamulin (Ribo-RET) to identify genome-wide translation start sites in the human pathogen Streptococcus pneumoniae. We identified 114 unannotated intergenic small open reading frames (sORFs). The described procedures and data sets provide a model for microbiologists seeking to explore the translational landscape of bacteria. The biological roles of four sORF examples are characterized: two control the regulation of a cell-cell communication (quorum sensing) system, one contributes to the ability of S. pneumoniae to colonize the upper respiratory tract of mice, and a fourth governs the translation of PrfB, a protein enabling ribosome release at stop codons. We propose that Ribo-RET is a valuable approach to identifying unstudied microproteins and difficult-to-find pheromone genes used by Gram-positive organisms, whose genomes are replete with pheromone receptors"
Keywords:		Animals Humans Mice Open Reading Frames *Quorum Sensing/genetics Ribosomes/genetics/metabolism *Streptococcus pneumoniae/genetics Virulence Streptococcus pneumoniae D39 quorum sensing ribosome profiling small open reading frames small proteins translation;
Notes:		"MedlineLaczkovich, Irina Mangano, Kyle Shao, Xinhao Hockenberry, Adam J Gao, Yu Mankin, Alexander Vazquez-Laslop, Nora Federle, Michael J eng R01 AI091779/AI/NIAID NIH HHS/ R21 AI144500/AI/NIAID NIH HHS/ Research Support, N.I.H., Extramural 2022/07/20 mBio. 2022 Aug 30; 13(4):e0124722. doi: 10.1128/mbio.01247-22. Epub 2022 Jul 19"