A computational and secretome analysis approach reveals exclusive and shared candidate type six secretion system substrates in Pectobacterium brasiliense 1692

Abstract

The type 6 secretion system (T6SS) of Gram-negative bacteria (GNB) has implications for bacterial competition, virulence, and survival. For the broad host range pathogen, Pectobacterium brasiliense 1692, T6SS-mediated competition occurs in a tissue-specific manner. However, no other roles have been investigated. The aim of this study was to identify T6SS-associated proteins under virulence inducing conditions. We used Bastion tools to predict 1479 Pbr1692 secreted proteins. Sixteen percent of these overlap between type 1–4 secretion systems (T1SS-T4SS) and T6SS. Using label-free quantitative mass spectrometry of Pbr1692 T6SS active and T6SS inactive strains’ secretomes cultured in minimal media supplemented with host extract, 49 T6SS-associated proteins with varied gene ontology predicted functions were identified. We report 19 and 30 T6SS primary substrates and differentially secreted proteins, respectively, in T6SS mutants versus wild type strains. Of the total 49 T6SS-associated proteins presented in this study, 25 were also predicted using the BastionX platform as T6SS exclusive and shared substrates with T1SS-T4SS. This work provides a list of Pbr1692 T6SS secreted effector candidates. These include a potential antibacterial toxin HNH endonuclease and several predicted virulence proteins, including plant cell wall degrading enzymes. A preliminary basis for potential crosstalk between GNB secretion systems is also highlighted.