Role of the sRNA GcvB in regulation of cycA in Escherichia coli

• Published in: Microbiology (Society for General Microbiology) Vol. 155; no. 1; pp. 106 - 114
• Main Authors: Pulvermacher, Sarah C, Stauffer, Lorraine T, Stauffer, George V
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.01.2009; Society for General Microbiology
• Subjects: Amino Acid Transport Systems - genetics; Amino Acid Transport Systems - metabolism; Bacteriology; Biological and medical sciences; Cycas; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli - physiology; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Bacterial; Genetics; Host Factor 1 Protein - genetics; Host Factor 1 Protein - metabolism; Microbiology; Mutagenesis, Site-Directed; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Bacterial - genetics; RNA, Bacterial - metabolism; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA, Untranslated - genetics; RNA, Untranslated - metabolism; Bacteria; Regulation(control); Physiology; RNA; Escherichia coli; Enterobacteriaceae
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/mic.0.023598-0

Department of Microbiology, University of Iowa, Iowa City, IA 52242, USA Correspondence George V. Stauffer george-stauffer{at}uiowa.edu In Escherichia coli , the gcvB gene encodes a small non-translated RNA that regulates several genes involved in transport of amino acids and peptides (including sstT , oppA and dppA ). Microarray analysis identified cycA as an additional regulatory target of GcvB. The cycA gene encodes a permease for the transport of glycine, D -alanine, D -serine and D -cycloserine. RT-PCR confirmed that GcvB and the Hfq protein negatively regulate cycA mRNA in cells grown in Luria–Bertani broth. In addition, deletion of the gcvB gene resulted in increased sensitivity to D -cycloserine, consistent with increased expression of cycA . A cycA : : lacZ translational fusion confirmed that GcvB negatively regulates cycA expression in Luria–Bertani broth and that Hfq is required for the GcvB effect. GcvB had no effect on cycA : : lacZ expression in glucose minimal medium supplemented with glycine. However, Hfq still negatively regulated the fusion in the absence of GcvB. A set of transcriptional fusions of cycA to lacZ identified a sequence in cycA necessary for regulation by GcvB. Analysis of GcvB identified a region complementary to this region of cycA mRNA. However, mutations predicted to disrupt base-pairing between cycA mRNA and GcvB did not alter expression of cycA : : lacZ . A model for GcvB function in cell physiology is discussed. Abbreviations: DEPC, diethylpyrocarbonate; sRNA, small non-translated regulatory RNA; WT, wild-type A supplementary table listing genes shown to be significantly regulated in the WT versus the gcvB strain by microarray analysis is available with the online version of this paper.