title: Regulatory network controlling extracellular proteins in Erwinia carotovora subsp. carotovora: FlhDC, the master regulator of flagellar genes, activates rsmB regulatory RNA production by affecting gacA and hexA (lrhA) expression.
abstract: Erwinia carotovora subsp. carotovora produces an array of extracellular proteins (i.e., exoproteins), including plant cell wall-degrading enzymes and Harpin, an effector responsible for eliciting hypersensitive reaction. Exoprotein genes are coregulated by the quorum-sensing signal, N-acyl homoserine lactone, plant signals, an assortment of transcriptional factors/regulators (GacS/A, ExpR1, ExpR2, KdgR, RpoS, HexA, and RsmC) and posttranscriptional regulators (RsmA, rsmB RNA). rsmB RNA production is positively regulated by GacS/A, a two-component system, and negatively regulated by HexA (PecT in Erwinia chrysanthemi; LrhA [LysR homolog A] in Escherichia coli) and RsmC, a putative transcriptional adaptor. While free RsmA, an RNA-binding protein, promotes decay of mRNAs of exoprotein genes, binding of RsmA with rsmB RNA neutralizes the RsmA effect. In the course of studies of GacA regulation, we discovered that a locus bearing strong homology to the flhDC operon of E. coli also controls extracellular enzyme production. A transposon insertion FlhDC(-) mutant produces very low levels of pectate lyase, polygalacturonase, cellulase, protease, and E. carotovora subsp. carotovora Harpin (Harpin(Ecc)) and is severely attenuated in its plant virulence. The production of these exoproteins is restored in the mutant carrying an FlhDC(+) plasmid. Sequence analysis and transcript assays disclosed that the flhD operon of E. carotovora subsp. carotovora, like those of other enterobacteria, consists of flhD and flhC. Complementation analysis revealed that the regulatory effect requires functions of both flhD and flhC products. The data presented here show that FlhDC positively regulates gacA, rsmC, and fliA and negatively regulates hexA (lrhA). Evidence shows that FlhDC controls extracellular protein production through cumulative effects on hexA and gacA. Reduced levels of GacA and elevated levels of HexA in the FlhDC(-) mutant are responsible for the inhibition of rsmB RNA production, a condition conducive to the accumulation of free RsmA. Indeed, studies with an RsmA(-) FlhDC(-) double mutant and multiple copies of rsmB(+) DNA establish that the negative effect of FlhDC deficiency is exerted via RsmA. The FlhDC-mediated regulation of fliA has no bearing on exoprotein production in E. carotovora subsp. carotovora. Our observations for the first time establish a regulatory connection between FlhDC, HexA, GacA, and rsmB RNA in the context of the exoprotein production and virulence of E. carotovora subsp. carotovora.
authors.affiliation: Division of Plant Sciences, 108 Waters Hall, University of Missouri, Columbia, MO 65211, USA.
authors.forename: Yaya
authors.initials: Y
authors.lastname: Cui
authors.forename: Asita
authors.initials: A
authors.lastname: Chatterjee
authors.forename: Hailian
authors.initials: H
authors.lastname: Yang
authors.forename: Arun K
authors.initials: AK
authors.lastname: Chatterjee
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015152:Blotting, Northern; D015153:Blotting, Western; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D008954:Models, Biological; D016973:Pectobacterium carotovorum / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism* / Q000502:physiology
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C064971:FliA protein, Bacteria; C091100:GacA protein, Bacteria; C084499:HexA protein, bacteria; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; C095468:RsmA protein, Erwinia carotovora; D012808:Sigma Factor; D014157:Transcription Factors; C115187:rsmb protein, Erwinia carotovora