Pseudomonas syringae pv. tomato DC3000 第六型分泌系統基因調控之初步探討

Due to the complexity of the double membrane structure, Gram-negative bacteria develop a variety of secretion systems, including Type I to Type V secretion system (T1SS to T5SS ) to secret proteins into the surrounding environments. In 2006, a novel secretion system, termed Type VI secretion system (T6SS), was recognized as another important virulence factor of the bacterial pathogen. In tomato pathogen, Pseudomonas syringae pv. tomato DC3000 (PstDC3000), two T6SS gene clusters in addition to its secretion components genes, hcp1 and hcp2, was identified using in silico analysis. Each cluster contains one copy of vasH homolog (PSPTO_2549 and PSPTO_5424), the gene coding for the sigma 54-dependent transcriptional activator, which controls the expression of T6SS-secreted protein, Hcp. By means of PSPTO_2549 and PSPTO_5424 mutants in accompany with a GUS reporter system, we showed that PSPTO_5424 plays a very important role in regulation of hcp-2 expression and PSPTO_2549 may cooperates with PSPTO_5424 to control the hcp-2 expression. Moreover, we also found that 240bp to 360bp upstream of the hcp-2 ORF is required for hcp-2 expression. Unlike type III secretion systems, genes in the second T6SS gene cluster or hcp-2 were expressed when PstDC3000 were grown in King’s B medium in a cell density-dependent manner, and expression of these genes is not completely induced by sugars as that of type III effector genes. The different expression patterns between T3SS and T6SS may explain why T6SS is not as important as T3SS in the virulence activity of PstDC3000 in planta.