An intrinsic mechanism for coordinated production of the contact-dependent and contact-independent weapon systems in a soil bacterium

Abstract

Soil bacteria possess multiple weapons to fend off microbial competitors. Currently, we poorly understand the factors guiding bacterial decisions about weapon systems deployment. In this study, we investigated how such decisions are made by the soil bacterium Lysobacter enzymogenes, used in antifungal plant protection. We found that weapons production is guided by environmental cues. In rich media, which likely mimic environments crowded with other microbes, L. enzymogenes produces a contact-dependent weapon, type six secretion system (T6SS). In nutrient-poor media, likely dominated by filamentous oomycetes and fungi, L. enzymogenes synthesizes and secretes a heat-stable antifungal factor (HSAF), a contact-independent weapon. Surprisingly, the T6SS inner tube protein Hcp is accumulated intracellularly even in nutrient-poor media, when the T6SS is not assembled. We found that Hcp interacts with the transcription factor Clp required for activating HSAF biosynthesis operon expression. Hcp protects Clp from binding to c-di-GMP, an intracellular second messenger inhibiting DNA binding. The increased concentration of c-di-GMP-free Clp thus leads to higher gene expression and HSAF production. Therefore, when the contact-dependent weapon, T6SS, is not in use, accumulation of one of its structural components, Hcp, serves as a signal to enhance production of the contact-independent weapon, HSAF. The uncovered environment-dependent and auto-regulatory mechanisms shed light on the processes governing deployment of various weapon systems in environmental bacteria.

Coordinated control of the type IV pili and c‐di‐GMP‐dependent antifungal antibiotic production in Lysobacter by the response regulator PilR

“…Protein purity was assessed by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE). Expression and purification of GST‐Clp, GST‐CdgL, and PilR‐His were described recently (Chen et al., 2017; Han et al., 2020; Xu et al., 2018; Yang, Ren, Shen, Yang, et al., 2020).…”

“…EMSA was performed as described previously (Ge et al., 2020; Yang, Ren, Shen, Yang, et al., 2020). The 5‐carboxy‐fluorescein (FAM)‐labelled probes comprising the promoter region upstream of lchD were synthesized by GENEWIZ.…”

“…Pull‐down assays were performed as described previously (Yang, Ren, Shen, Yang, et al., 2020). The reaction mixtures contained 5 μM of LchD‐His and Clp‐GST or CdgL‐GST in 1 ml of phosphate‐buffered saline (PBS).…”

Clp is a “busy” transcription factor in the bacterial warrior, Lysobacter enzymogenes

“…When a cell is in contact with another cell, the developed T6SS will pierce the cell wall and cell membrane of the prey (target) cell, and transmits a toxin-laden shell (known as the inner tube) to it [66] , [67] . L. enzymogenes also possesses a complete T6SS gene cluster that can work together to assemble a functional T6SS, as evidenced by the secretion of Hcp when L. enzymogenes is cultured in a nutrient-rich medium (LB) that likely mimics an environment rich in other microorganisms [68] . However, under the same nutritional conditions, the production of “long-range” weapon is hindered [68] .…”

“…L. enzymogenes also possesses a complete T6SS gene cluster that can work together to assemble a functional T6SS, as evidenced by the secretion of Hcp when L. enzymogenes is cultured in a nutrient-rich medium (LB) that likely mimics an environment rich in other microorganisms [68] . However, under the same nutritional conditions, the production of “long-range” weapon is hindered [68] . In contrast, in the HSAF-producing medium (1/10 TSB), which is likely to be present in filamentous oomycetes and fungi, T6SS is not assembled and Hcp secretion is not detected [68] .…”