FleQ regulates both the type VI secretion system and flagella in Pseudomonas putida

Based on the analysis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and mass spectrometry, flagellin FliC and a key member of the type VI secretion system Hcp1 were found to be secreted from Pseudomonas putida KT2442 cells in the late exponential phase, but in the early exponential phase only FliC was secreted. Relevant genes fleQ and rpoN were then individually deleted in P. putida KT2442, and the expression and transcription of fliC and hcp1 in these mutant strains were analyzed. In both △fleQ and △rpoN mutant cells, Hcp1 but not FliC was secreted in the late exponential phase, and it was secreted even in the early exponential phase. Transcriptomic analysis of △fleQ cells shows that most of the genes relevant to the type VI secretion system, including hcp1, were upregulated; several genes involved in 3',5'-cyclic diguanylic acid (c-di-GMP) synthesis and degradation were also significantly regulated. To further study the regulation process of the type VI secretion system, the phosphodiesterase BifA was overexpressed to lower the intracellular c-di-GMP level. The results demonstrate that FleQ positively regulates flagella biosynthesis but negatively regulates the type VI secretion system in P. putida. This finding is important for understanding the function and regulation of the type VI secretion system.