Genome‐wide identification of genes directly regulated by ChvI and a consensus sequence for ChvI binding in Sinorhizobium meliloti

Summary ExoS/ChvI two‐component signaling in the nitrogen‐fixing α‐proteobacterium Sinorhizobium meliloti is required for symbiosis and regulates exopolysaccharide production, motility, cell envelope integrity and nutrient utilization in free‐living bacteria. However, identification of many ExoS/Chv...

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Vydáno v:	Molecular microbiology Ročník 110; číslo 4; s. 596 - 615
Hlavní autoři:	Ratib, Nicole R., Sabio, Erich Y., Mendoza, Carolina, Barnett, Melanie J., Clover, Sarah B., Ortega, Jesus A., Dela Cruz, Francesca M., Balderas, David, White, Holly, Long, Sharon R., Chen, Esther J.
Médium:	Journal Article
Jazyk:	angličtina
Vydáno:	England Blackwell Publishing Ltd 01.11.2018
Témata:	Bacterial Proteins - genetics Bacterial Proteins - metabolism Binding Binding Sites - genetics Chromatin Conserved sequence Deoxyribonucleic acid DNA DNA microarrays DNA-Binding Proteins - genetics Electrophoretic mobility Exopolysaccharides Gene expression Gene Expression Regulation, Bacterial - genetics Gene sequencing Genes Genome, Bacterial - genetics Genomes Glucosyltransferases - genetics Immunoprecipitation Nucleotide sequence Nutrient utilization Protein Binding - genetics Proteins Regulators Signal Transduction Sinorhizobium meliloti Sinorhizobium meliloti - genetics Sinorhizobium meliloti - metabolism Symbiosis Symbiosis - genetics Target recognition Transcription Transcription Factors - genetics Transcription Factors - metabolism Transcription, Genetic - genetics
ISSN:	0950-382X, 1365-2958, 1365-2958
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Shrnutí:	Summary ExoS/ChvI two‐component signaling in the nitrogen‐fixing α‐proteobacterium Sinorhizobium meliloti is required for symbiosis and regulates exopolysaccharide production, motility, cell envelope integrity and nutrient utilization in free‐living bacteria. However, identification of many ExoS/ChvI direct transcriptional target genes has remained elusive. Here, we performed chromatin immunoprecipitation followed by microarray analysis (chIP‐chip) to globally identify DNA regions bound by ChvI protein in S. meliloti. We then performed qRT‐PCR with chvI mutant strains to test ChvI‐dependent expression of genes downstream of the ChvI‐bound DNA regions. We identified 64 direct target genes of ChvI, including exoY, rem and chvI itself. We also identified ChvI direct target candidates, like exoR, that are likely controlled by additional regulators. Analysis of upstream sequences from the 64 ChvI direct target genes identified a 15 bp‐long consensus sequence. Using electrophoretic mobility shift assays and transcriptional fusions with exoY, SMb21440, SMc00084, SMc01580, chvI, and ropB1, we demonstrated this consensus sequence is important for ChvI binding to DNA and transcription of ChvI direct target genes. Thus, we have comprehensively identified ChvI regulon genes and a ‘ChvI box’ bound by ChvI. Many ChvI direct target genes may influence the cell envelope, consistent with the critical role of ExoS/ChvI in growth and microbe–host interactions. The Sinorhizobium meliloti ExoS/ChvI two‐component signaling pathway is critical for establishing a nitrogen‐fixing symbiosis with plant hosts and regulates many free‐living bacterial phenotypes. We report the comprehensive identification of genes comprising the ChvI regulon in S. meliloti and demonstrate the importance of a consensus sequence for ChvI‐DNA binding and regulation of ChvI direct target genes. These findings represent a major advance in understanding the role of this conserved signaling pathway involved in successful microbe–host interactions.
Bibliografie:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 14 content type line 23 Author Contributions EYS, NRR, CM, MJB, and EJC conceived and designed the study; NRR, EYS, CM, MJB, SBC, JAO, FMDC, DB, HW, and EJC acquired, analyzed, and interpreted data; and MJB, SRL, NRR, EYS, and EJC wrote the manuscript.
ISSN:	0950-382X 1365-2958 1365-2958
DOI:	10.1111/mmi.14119