Identification of regulators of hrp / hop genes of Erwinia carotovora ssp. carotovora and characterization of HrpL Ecc (SigmaL Ecc ), an alternative sigma factor

Summary Erwinia carotovora ssp. carotovora (Ecc) possesses hrpN Ecc [ hrp  = gene for hypersensitive reaction (HR) and pathogenicity], the structural gene for Harpin Ecc , the inducer of the HR‐like response and genes for the type III secretion system. In Ecc, RsmA, an RNA‐binding protein responsible for the accelerated decay of RNA species, tightly controls the expression of the Hrp regulon. We document here that Ecc strain 71 possesses several presumed Hrp regulators: HrpX Ecc , HrpY Ecc , HrpS Ecc , and HrpL Ecc . Nucleotide sequence data indicate that the regulatory genes occur as a cluster. The data also suggest that HrpX Ecc is a putative sensor kinase, HrpY Ecc is the cognate response regulator, and HrpS Ecc is an NtrC‐like regulator. hrpL Ecc consists of a 543‐bp open reading frame (ORF), which encodes a c . 21‐kDa protein product. This protein shares significant homology with members of the extracytoplasmic function (ECF) subfamily of alternative sigma factors, including SigE. To examine the regulatory role of hrpL Ecc , we constructed by marker exchange an HrpL Ecc − derivative of an RsmA − strain. The levels of transcripts of hrcC Ecc , a gene for the type III secretion system, and hrpN Ecc were lower in the RsmA − HrpL Ecc − mutant compared to its RsmA − HrpL Ecc + parent. The RsmA − HrpL Ecc − strain, like the RsmA − HrpL Ecc + parent, caused maceration of celery petioles and produced extracellular pectinases, cellulase, and protease, indicating that the alternative sigma factor is not required for the type I and type II secretion systems, for tissue maceration, or for the production of proteins secreted by these pathways. However, the RsmA − HrpL Ecc − strain, unlike the RsmA − HrpL Ecc + strain, did not elicit the HR in tobacco leaves. In addition, the RsmA − HrpL − strain failed to cause symptoms in Arabidopsis thaliana . Genetic and biochemical data revealed that Harpin Ecc is required for symptom production in Arabidopsis , but is not sufficient by itself to cause necrosis. These observations raise the possibility that another ‘effector’ protein of Ecc, secreted by the type III system, acts in conjunction with Harpin Ecc to trigger responses resulting in cell death in Arabidopsis .