Journal
MICROBIOLOGY SPECTRUM
Volume 10, Issue 4, Pages -Publisher
AMER SOC MICROBIOLOGY
DOI: 10.1128/spectrum.01576-22
Keywords
type VI secretion system; VgrG proteins; interbacterial competition; Chromobacterium violaceum; quorum sensing
Categories
Funding
- Sao Paulo Research Foundation (FAPESP) [2020/00259-8, 2021/06894-0]
- Fundacao de Apoio ao Ensino, Pesquisa e Assistencia do Hospital das Clinicas da FMRP-USP (FAEPA)
- FAPESP [2018/03979-1, 2018/12461-6]
- Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq)
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The environmental bacterium Chromobacterium violaceum uses a type VI secretion system (T6SS) to compete with other bacteria, and this competition mechanism is independent of its infection in mice. VgrG proteins play key roles in the T6SS function, and different VgrGs show little functional redundancy. The expression of T6SS genes is regulated by cell density, and the regulation is dependent on CviR.
The environmental pathogenic bacterium Chromobacterium violaceum kills Gram-positive bacteria by delivering violacein packed into outer membrane vesicles, but nothing is known about its contact-dependent competition mechanisms. In this work, we demonstrate that C. violaceum utilizes a type VI secretion system (T6SS) containing multiple VgrG proteins primarily for interbacterial competition. The single T6SS of C. violaceum contains six vgrG genes, which are located in the main T6SS cluster and four vgrG islands. Using T6SS core component-null mutant strains, Western blotting, fluorescence microscopy, and competition assays, we showed that the C. violaceum T6SS is active and required for competition against Gram-negative bacteria such as Pseudomonas aeruginosa but dispensable for C. violaceum infection in mice. Characterization of single and multiple vgrG mutants revealed that, despite having high sequence similarity, the six VgrGs show little functional redundancy, with VgrG3 showing a major role in T6SS function. Our coimmunoprecipitation data support a model of VgrG3 interacting directly with the other VgrGs. Moreover, we determined that the promoter activities of T6SS genes increased at high cell density, but the produced Hcp protein was not secreted under such condition. This T6SS growth phase-dependent regulation was dependent on CviR but not on CviI, the components of a C. violaceum quorum sensing (QS) system. Indeed, a DcviR but not a DcviI mutant was completely defective in Hcp secretion, T6SS activity, and interbacterial competition. Overall, our data reveal that C. violaceum relies on a QS-regulated T6SS to outcompete other bacteria and expand our knowledge about the redundancy of multiple VgrGs. IMPORTANCE The type VI secretion system (T6SS) is a contractile nanomachine used by many Gram-negative bacteria to inject toxic effectors into adjacent cells. The delivered effectors are bound to the components of a puncturing apparatus containing the protein VgrG. The T6SS has been implicated in pathogenesis and, more commonly, in competition among bacteria. Chromobacterium violaceum is an environmental bacterium that causes deadly infections in humans. In this work, we characterized the single T6SS of C. violaceum ATCC 12472, including its six VgrG proteins, regarding its function and regulation. This previously undescribed C. violaceum T6SS is active, regulated by QS, and required for interbacterial competition instead of acute infection in mice. Among the VgrGs, VgrG3, encoded outside the main T6SS cluster, showed a major contribution to T6SS function. These results shed light on a key contact-dependent killing mechanism used by C. violaceum to antagonize other bacteria.
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