Novel polyadenylylation-dependent neutralization mechanism of the HEPN/MNT toxin/antitoxin system

Abstract

The two-gene module HEPN/MNT is predicted to be the most abundant toxin/antitoxin (TA) system in prokaryotes. However, its physiological function and neutralization mechanism remains obscure. Here, we discovered that the MntA antitoxin (MNT-domain protein) acts as an adenylyltransferase and chemically modifies the HepT toxin (HEPN-domain protein) to block its toxicity as an RNase. Biochemical and structural studies revealed that MntA mediates the transfer of three AMPs to a tyrosine residue next to the RNase domain of HepT in Shewanella oneidensis. Furthermore, in vitro enzymatic assays showed that the three AMPs are transferred to HepT by MntA consecutively with ATP serving as the substrate, and this polyadenylylation is crucial for reducing HepT toxicity. Additionally, the GSX10DXD motif, which is conserved among MntA proteins, is the key active motif for polyadenylylating and neutralizing HepT. Thus, HepT/MntA represents a new type of TA system, and the polyadenylylation-dependent TA neutralization mechanism is prevalent in bacteria and archaea.

Identification of a Toxin–Antitoxin System That Contributes to Persister Formation by Reducing NAD in Pseudomonas aeruginosa

“…Based on the natures of the antitoxins and their mechanisms of action, the TA systems have been classified into six types (I–VI) [ 9 ]. Recently, a group of type VII TA systems was discovered in Escherichia coli [ 14 ], Yersinia enterocolitica [ 15 ], Mycobacterium tuberculosis [ 16 ] and Shewanella oneidensis [ 17 , 18 ]. The type II TA systems are widespread in bacterial genomes [ 19 ].…”

HEPN-MNT Toxin-Antitoxin System: The HEPN Ribonuclease Is Neutralized by OligoAMPylation

“…S. oneidensis MNT and HEPN proteins form a hetero-octameric complex with a MNT 2 -HEPN 6 stoichiometry (Jia et al, 2018). While this paper was under consideration, a paper providing further biochemical and structural characterization of S. oneidensis HEPN-MNT system appeared (Yao et al, 2020).…”

Prokaryote toxin–antitoxin modules: Complex regulation of an unclear function

“…A second example of inactivation via posttranslational modification is the poly-adenylation found in the mntA-hepT family. 47,48 In this case, binding of antitoxin MntA is insufficient to fully inhibit HepT toxicity and the antitoxin MntA additionally functions as an adenylyltransferase. The toxic activity of the HepT RNase is neutralized by MntA via modification of a tyrosine residue close to the toxin's active site.…”

“…A second example of inactivation via post‐translational modification is the poly‐adenylation found in the mntA‐hepT family 47,48 . In this case, binding of antitoxin MntA is insufficient to fully inhibit HepT toxicity and the antitoxin MntA additionally functions as an adenylyltransferase.…”