Abstract
Type VI secretion systems (T6SS) are multiprotein secretion machines that can mediate killing of bacterial cells and thereby modify the composition of bacterial communities. The mechanisms that control the production of and secretion of these killing machines are incompletely understood, although quorum sensing (QS) and the PpkA kinase modulate T6SS activity in some organisms. Here we investigated control the T6S in the marine organism Vibrio alginolyticus EPGS, which encodes two T6SS systems (T6SS1 and T6SS2). We found that the organism principally relies on T6SS2 for interbacterial competition. We further carried out a phosphoproteomic screen to identify substrates of the T6SS2-linked PpkA2 kinase. Substrates of PpkA2 encoded within the T6SS2 cluster as well proteins that are apparently not linked to T6SS-related processes were identified. Similar to other organisms, PpkA2 autophosphorylation was critical for T6SS2 function. Notably, phosphorylation of a polypeptide encoded outside of the T6SS2 cluster, VtsR, was critical for T6SS2 expression and function because it augments the expression of luxR, a key regulator of QS that also promotes T6SS2 gene expression. Thus, PpkA2 controls a phosphorylation cascade that mediates a positive regulatory loop entwining T6SS and QS, thereby coordinating these pathways to enhance the competitive fitness of V. alginolyticus.