Manipulation of GameXPeptide synthetase gene expression by a promoter exchange alters the virulence of an entomopathogenic bacterium, , by modulating insect immune responses.

An entomopathogenic bacterium, subsp. , is mutualistic to its host nematode, . The infective juvenile nematodes enter target insects through natural openings and release the symbiotic bacteria into the insect hemocoel. The released bacteria suppress the insect immune responses and cause septicemia through their secondary metabolites. GameXPeptide (GXP) is one of the common secondary metabolites of most species and is produced by the catalytic activity of a specific non-ribosomal peptide synthetase called GxpS encoded by the gene. This study confirmed to be encoded in the genome and analyzed its expression during bacterial growth. LC-MS/MS analysis of the bacterial culture broth contained at least four different GXPs (GXP-A to GXP-D), in which GXP-A was the most abundant. To investigate GXP synthesis following expression, the promoter of was replaced with an inducible arabinose promoter by homologous recombination. The transcript levels in the mutant were altered by the addition of l-arabinose. Without the inducer, the transcript level was significantly lower compared to the wild type and produced significantly lower amounts of the four GXPs. The addition of the inducer to the mutant significantly increased expression and produced significantly higher levels of the four GXPs compared to the wild type. The metabolite extracts obtained from wild-type and mutant bacteria showed differential immunosuppressive activities according to their GXP contents against the cellular and humoral immune responses of a lepidopteran insect, . Interestingly, the -mutant bacteria showed less insecticidal activity compared to the wild type, whereas the addition of GXP to the mutant significantly restored insecticidal activity. These results suggest that the gene encoded in is responsible for the production of at least four different GXPs, which play crucial roles in bacterial virulence.