An interbacterial lipase toxin with an unprecedented reverse domain arrangement defines a new class of type VII secretion system effector

Abstract

The type VII protein secretion system (T7SS) is found in many Gram-positive bacteria and in pathogenic mycobacteria. All T7SS substrate proteins described to date share a common helical domain architecture at the N-terminus that typically interacts with other helical partner proteins, forming a composite signal sequence for targeting to the T7SS. The C-terminal domains are functionally diverse and in Gram-positive bacteria such as Staphylococcus aureus often specify toxic anti-bacterial activity. Here we describe the first example of a new class of T7 substrate, TslA, that has an unexpected reverse domain organisation. TslA is widely found across Bacillota including Staphylococcus, Enterococcus and Listeria. We show that the S. aureus TslA N-terminal domain is a phospholipase A with anti-staphylococcal activity that is neutralised by the immunity lipoprotein TilA. Two small helical partner proteins, TlaA1 and TlaA2 are essential for T7-dependent secretion of TslA and at least one of these interacts with the TslA C-terminal domain to form a helical stack. Cryo-EM analysis of purified TslA complexes indicate that they share structural similarity with canonical T7 substrates. Our findings suggest that the T7SS has the extraordinary feature of recognising a secretion signal present at either end of a substrate.

Interbacterial competition mediated by the type VIIb secretion system

“…esxB , esxC and esxD are located in a four-gene cluster along with esaE , directly upstream of esaD (), a synteny which is conserved among esaD / yeeF homologues. Similarly, secretion of the reverse substrate TslA also utilises a WXG100-like pair of export factors, Tla1 and Tla2, which are encoded immediately downstream of the toxin, and likely bind analogously to its reverse-LXG domain [73].…”

“…The targeting domain is not simply appended to the C-terminus of the protein, but the entire sequence is reversed and so carries a GxL motif. These proteins, named TslA ( T ype VII s ecreted l ipase A), are distributed widely among Bacillota and are also verified T7SSb substrates [ 73 ].…”

“…TslA carries a C-terminal GXL targeting domain, and the N-terminal toxic domain shows phospholipase A activity. It is neutralised though tight interaction with its cognate immunity protein, TilA [ 73 ].…”