No abstract

Gram-positive bacteria deploy the type VII secretion system (T7SS) to facilitate interactions between eukaryotic and prokaryotic cells. In recent work, we identified the TelC protein from Streptococcus intermedius as a T7SS-exported lipid II phosphatase that mediates interbacterial competition. TelC exerts toxicity in the inner wall zone of Gram-positive bacteria; however, intercellular intoxication of sister cells does not occur because they express the TipC immunity protein. In the present study, we sought to characterize the molecular basis of self-protection by TipC. Using sub-cellular localization and protease protection assays, we show that TipC is a membrane protein with an N-terminal transmembrane segment and a C-terminal TelC-inhibitory domain that protrudes into the inner wall zone. The 1.9-Å X-ray crystal structure of a non-protective TipC paralogue reveals that the soluble domain of TipC proteins adopts a crescent-shaped fold that is composed of three α-helices and a seven-stranded β-sheet. Subsequent homology-guided mutagenesis demonstrates that a concave surface formed by the predicted β-sheet of TipC is required for both its interaction with TelC and its TelC-inhibitory activity. S. intermedius cells lacking the tipC gene are susceptible to growth inhibition by TelC delivered between cells; however, we find that the growth of this strain is unaffected by endogenous or overexpressed TelC, although the toxin accumulates in culture supernatants. Together, these data indicate that the TelC-inhibitory activity of TipC is only required for intercellularly transferred TelC and that the T7SS apparatus transports TelC across the cell envelope in a single step, bypassing the cellular compartment in which it exerts toxicity en route.

Type VIIb secretion systems (T7SSb) are protein secretion machines used by an array of Gram-positive bacterial genera, including Staphylococcus , Streptococcus , Bacillus , and Enterococcus . These bacteria use the T7SSb to facilitate interbacterial killing and pathogenesis through the secretion of toxins.

Type VII secretion systems (T7SSs) are poorly understood protein export apparatuses found in mycobacteria and many species of Gram-positive bacteria. To date, this pathway has predominantly been studied in Mycobacterium tuberculosis, where it has been shown to play an essential role in virulence; however, much less studied is an evolutionarily divergent subfamily of T7SSs referred to as the T7SSb. The T7SSb is found in the major Gram-positive phylum Firmicutes where it was recently shown to target both eukaryotic and prokaryotic cells, suggesting a dual role for this pathway in host-microbe and microbe-microbe interactions. In this review, we compare the current understanding of the molecular architectures and substrate repertoires of the well-studied mycobacterial T7SSa systems to that of recently characterized T7SSb pathways and highlight how these differences may explain the observed biological functions of this understudied protein export machine.

Highlights d EsaA is an essential component of the Streptococcus intermedius T7SSb d Topology experiments suggest a large soluble domain of EsaA is extracellular d Structure of EsaA extracellular region reveals a dimer comprised of three domains d EsaA may form a conduit that facilitates effector secretion across the cell wall