A novel seven-helix transmembrane protein BTP1 of Botrytis cinerea controls the expression of GST-encoding genes, but is not essential for pathogenicity.

SUMMARY To gain new insights into the signalling mechanisms of the grey mould Botrytis cinerea, which causes several pre- and post-harvest diseases on a variety of host plants, we cloned, sequenced and functionally characterized a gene, btp1, encoding a novel 391-amino acid transmembrane protein. The protein BTP1 shows similarity to the transmembrane protein pth11, which is essential for appressorium formation and successful colonization of plant tissue in the rice blast fungus Magnaporthe grisea. Analyses of the deduced amino acid sequence of btp1 predicted a seven alpha-helical transmembrane topology, which is known to be typical for G protein-coupled receptors (GPCRs) and therefore the protein is thought to play a role in mediation of extracellular signals to intracellular effectors. The gene is located next to the gene bcgstII encoding a new putative glutathione S-transferase, and both genes are transcribed in opposite directions from the same promoter. BcGSTII shows similarity to the glutathione S-transferase GSTII of Schizosaccharomyces pombe, a protein thought to be involved in detoxification of several antifungal drugs. From the sequence similarity of BTP1 to GPCRs, and its expression in planta, we suggested that it might play a role in mediation of plant signals and therefore in pathogenicity. However, targeted gene replacement of btp1 did not result in a phenotype markedly affecting either pathogenicity or sensitivity to chemical stress when compared with the wild-type strain; however, the ten-fold dilution of conidial suspension used for the pathogenicity assay resulted in slight reduction of virulence. Visible symptom development of the mutants on bean plants was also different from the wild-type. The brownish ring, which appears at the margin of secondary lesions in wild-type infections, was brighter and almost absent in Deltabtp1 mutants. Interestingly, deletion of btp1 not only affected the expression of the physically linked bcgstII gene, but in addition the expression of the other two GST-encoding genes in B. cinerea for bcgstI was down-regulated, bcgstII was slightly up-regulated and bcgstIII was strongly up-regulated in the mutant.