Acidovorax citrulli Type IV Pili PilR Interacts with PilS and Regulates the Expression of the pilA Gene

Abstract

Acidovorax citrulli can cause bacterial fruit blotch of watermelon, melon, and other cucurbits, and has the potential to cause severe economic losses to growers throughout the world. This article investigated the functions and interactions of the pilR and pilS genes, two important genes in bacterial type IV pili systems, in A. citrulli. For each gene, deletion mutants and complementary strains were constructed via homologous recombination, and their phenotypes were determined. The results showed that the absence of pilR and pilS could significantly reduce the pathogenicity and twitching motility of A. citrulli while increasing the swimming motility, biofilm formation, and in vitro growth. Conversely, complementary strains were no different than the wild-type strain. Using quantitative reverse transcription PCR and promoter activity assays, we confirmed that the deletion of pilR and pilS genes leads to a significant decrease in the transcription level of pilA. Meanwhile, three methods including yeast two-hybrid, glutathione S-transferase pull-down, and luciferase complementation imaging assays were used to verify the direct interaction between the PilR and PilS proteins. These findings revealed the biological function of the pilR and pilS and confirms their regulatory role on pilA.