Identification of a putative alternate sigma factor and characterization of a multicomponent regulatory cascade controlling the expression of Pseudomonas syringae pv. syringae Pss61 hrp and hrmA genes

Abstract

The Pseudomonas syringae hrp and hrmA genes controlling pathogenicity and elicitation of the hypersensitive response and the avr genes controlling host range have been shown previously to be regulated by carbon, nitrogen, pH, osmolarity, and hypothetical plant factors. In P. syringae pv. syringae Pss6l, inactivation of hrp complementation groups II and XIII reduced expression of a plasmid-borne hrmA'-lacZ fusion. The hrp regions II and XIII were cloned on separate plasmids and shown to enhance the activity of the hrmA promoter in Escherichia coli MC4100 transformants at least 100-fold. The nucleotide sequence of region XIII revealed two open reading frames (hrpR and hrpS) whose deduced products share homology with P. syringae pv. phaseolicola NPS3121 HrpS and are both related to the NtrC family of two-component signal transduction systems. HrpR and HrpS differ from most members of the protein family by lacking an amino-terminal domain which modulates the regulatory activity. A single open reading frame, hrpL, whose product shares homology with AlgU, a putative alternate sigma factor of P. aeruginosa, as well as with the related alternate sigma factors was identified. within region II. Key domains are partially conserved. Inactivation of hrpS in Pss6l repressed expression of a plasmid-borne hrpL'-lacZ fusion carried by pYXPLlR, and transformation of MC4100(pYXPL1R) with a plasmid carrying hrpRS increased hrpL promoter activity at least 200-fold. Neither hrpS nor hrpR, when cloned on separate plasmids, activated the hrpL promoter activity individually. The expression of hrpL when directed by a lac promoter was sufficient to express a set of plasmid-borne hrmA'-, hrpJ'-, and hrpZ'-lacZ fusions independently of other hrp genes. The results indicate that hrpRS and hrpL are part of a regulatory cascade in which HrpR and HrpS activate expression of hrpL and HrpL, a putative sigma factor, induces expression of HrpL-responsive genes.

The Escherichia coli phage‐shock‐protein (psp) operon

“…PspF lacks an N-terminal domain and is constitutively active in vivo and in vitro, consistent with the observation that deletion of the pspA gene results in high-level, constitutive expression from the psp promoter. The constitutive activity of PspF is in accord with results from other laboratories, which have shown that deletion of the N-terminal part of some other 54 activators leaves them constitutively active (Grimm et al, 1995;Gu et al, 1994;Huala et al, 1992;Lee et al, 1994;PerezMartin and de Lorenzo, 1996;Ronson, 1988;Xiao et al, 1994).…”

“…In contrast to the other genes of the psp cluster, pspF is related to many others in the database, and its product, PspF, is a member of the enhancer-binding-protein family of activators (Jovanovic et al, 1996). Most other such activators consist of an N-terminal domain that serves a regulatory function, a central catalytic domain required for activation which interacts with 54 -containing RNA polymerase, and a C-terminal domain that contains the helixturn-helix motif that constitutes a DNA-binding domain (Albright et al, 1989;Collado-Vides et al, 1991;Morett and Segovia, 1993;Parkinson and Kofoid, 1992;Shingler, 1996;Xiao et al, 1994). PspF lacks an N-terminal domain and is constitutively active in vivo and in vitro, consistent with the observation that deletion of the pspA gene results in high-level, constitutive expression from the psp promoter.…”

Virulence of the Phytopathogen Pseudomonas syringae pv. Maculicola Is rpoN Dependent

“…The hrpR product activates hrpS expression, and the hrpS product activates hrpL transcription (17,18). HrpL is an alternate sigma factor homologous to AlgU of P. aeruginosa and is thought to activate transcription of the remaining genes in the hrp cluster (96,97). The factor(s) involved in the regulation of hrpRS remains obscure.…”

Hrp pilus: An hrp -dependent bacterial surface appendage produced by Pseudomonas syringae pv. tomato DC3000

“…HrpL recognizes a consensus sequence motif (''harp box'') that has been identified in the upstream regions of many hrp and avr genes (4). The expression of hrp genes of many P. syringae pathovars can also be induced in vitro when bacteria are grown in defined minimal medium with low pH and containing certain sugars or sugar alcohols as carbon sources (5)(6)(7).…”

“…hrp genes of Pseudomonas syringae are expressed in planta as a result of a regulatory cascade involving the gene products of hrpS and hrpR, positive transcriptional regulators, and of hrpL, an alternative sigma factor (3,4). HrpL recognizes a consensus sequence motif (''harp box'') that has been identified in the upstream regions of many hrp and avr genes (4).…”