Inactivation of a gene that is highly conserved in Gram-positive bacteria stimulates degradation of non-native proteins and concomitantly increases stress tolerance in Lactococcus lactis.
Exposure of cells to elevated temperatures triggers the synthesis of chaperones and proteases including components of the conserved Clp protease complex. We demonstrated previously that the proteolytic subunit, ClpP, plays a major role in stress tolerance and in the degradation of non-native proteins in the Gram-positive bacterium Lactococcus lactis. Here, we used transposon mutagenesis to generate mutants in which the temperature- and puromycin-sensitive phenotype of a lactococcal clpP null mutant was partly alleviated. In all mutants obtained, the transposon was inserted in the L. lactis trmA gene. When analysing a clpP, trmA double mutant, we found that the expression normally induced from the clpP and dnaK promoters in the clpP mutant was reduced to wild-type level upon introduction of the trmA disruption. Additionally, the degradation of puromycyl-containing polypeptides was increased, suggesting that inactivation of trmA compensates for the absence of ClpP by stimulating an as yet unidentified protease that degrades misfolded proteins. When trmA was disrupted in wild-type cells, both stress tolerance and proteolysis of puromycyl peptides was enhanced above wild-type level. Based on our results, we propose that TrmA, which is well conserved in several Gram-positive bacteria, affects the degradation of non-native proteins and thereby controls stress tolerance.
Frees
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Department of Veterinary Microbiology, Stigbøjlen 4, Denmark.
Varmanen
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Ingmer
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D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004251:DNA Transposable Elements / Q000235:genetics; D049071:Endopeptidase Clp; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D018869:Heat-Shock Response; D013294:Lactococcus lactis / Q000187:drug effects / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016254:Mutagenesis, Insertional; D011489:Protein Denaturation; D017510:Protein Folding; D011691:Puromycin / Q000494:pharmacology; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D014158:Transcription, Genetic; D012359:tRNA Methyltransferases / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D011691:Puromycin; D012359:tRNA Methyltransferases; C022769:tRNA(uracil-5)-methyltransferase; D012697:Serine Endopeptidases; D049071:Endopeptidase Clp; D000251:Adenosine Triphosphatases