| The initial goal of the work presented here was to elucidate the mechanism of action of the elusive prlF1 suppressor. The prlF1 mutation was isolated as a suppressor of the toxicity of the hybrid protein, LamB-LacZ Hyb42-1, which causes a general, lethal, protein export defect, termed hybrid jamming, when its expression is induced. Mutations in the same gene were identified independently as suppressors of the temperature sensitive requirement for the periplasmic protease DegP (HtrA). Thus, prlF1 links two fields of long-standing interest: protein export, and envelope stress.; We knew from previous work that the prlF1 mutation activates the cytoplasmic protease Lon, and that a wildtype copy of the ton gene was required for all the phenotypes of prlF1. What was not clear was the nature of the interaction of the two genes, nor indeed what function the Lon protease served in the process, since there is no detectable increase in the proteolysis of the hybrid in prlF1 strains.; The work presented here falls broadly into two section. The first half describes a directed mutagenesis approach, which has clarified the recessive nature of the prlF1 suppressor, and which has led to believe that the mutant PrlF1 protein interacts directly with Lon. In the second half, I present the results of a screen for mutations that suppress the requirement for Ion, and in particular, the characterization of the ygdP gene. Two independent insertions in ygdP, which encodes a dinucleotide oligophosphate hydrolase, revela a novel mechanism of phenocopying prlF1. We propose two alternative models to explain the suppression of hybrid jamming by the loss of ygdP function. The remarkable commonality of the prlF1 and ygdP phenotypes suggests a fundamental link between protein export defects, and the requirement for degP at high temperatures. |
