Temperature regulation of protease in Pseudomonas fluorescens LS107d2 by an ECF sigma factor and a transmembrane activator

The production of extracellular enzymes by Pseudomonas fluorescens is impor tant with respect to phytopathogenesis and, in the case of psychrotrophic s trains, food spoilage. The production of extracellular protease has been pr eviously reported to be dependent on temperature in psychrotrophic strains of P. fluorescens; production is decreased above the optimum growth tempera ture with a relatively small change in growth rate. In this work, a transpo son mutant of P. fluorescens LS107d2 has been isolated which, in contrast t o the wild-type strain, is completely protease deficient at 29 degreesC, ab ove the optimum growth temperature of 25 degreesC, but which produces prote ase at 23 degreesC. Further analysis revealed that this mutation is in a ge ne (prtR) which is part of a dicistronic operon, prtIR, in which the two ge nes are translationally coupled. Evidence is presented that prtI encodes a sigma factor related to others involved in extracytoplasmic functions (ECF sigma factors) and that prtR encodes a novel transmembrane activator of Prt I. PrtI, like PrtR, is also required for protease production at 29 degreesC but not at 23 degreesC. Analysis of the amino acid sequence of PrtR indica tes that it is functionally related to a group of membrane-associated anti- sigma factors and a few transmembrane regulators, but is not significantly sequence related. Complementation analysis indicates that PrtR may also int eract with sigma factors other than PrtI. The promoter region of the protea se-encoding gene (aprX) in LS107d2 has been identified and has sequence fea tures which could indicate interaction with either an ECF sigma factor or a primary sigma factor.