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Novel polyadenylylation-dependent neutralization mechanism of the HEPN/MNT toxin/antitoxin system
Jianyun Yao1,2, Xiangkai Zhen3, Kaihao Tang1,2
1Key Laboratory of Tropical Marine Bio-resources and Ecology, Guangdong Key Laboratory of Marine Materia Medica, Innovation Academy of South China Sea Ecology and Environmental Engineering, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
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.
Insights
The HEPN/MNT toxin/antitoxin system uses polyadenylylation to neutralize toxicity. MntA chemically modifies HepT by adding AMPs, blocking its RNase activity and revealing a new neutralization mechanism in prokaryotes.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- The HEPN/MNT module is a prevalent toxin/antitoxin system in prokaryotes.
- Its precise function and neutralization mechanism are not well understood.
Purpose of the Study:
- To elucidate the physiological function and neutralization mechanism of the HEPN/MNT toxin/antitoxin system.
- To characterize the interaction between HepT toxin and MntA antitoxin.
Main Methods:
- Biochemical assays
- Structural studies
- In vitro enzymatic assays
- Site-directed mutagenesis
Main Results:
- MntA acts as an adenylyltransferase, chemically modifying the HepT toxin.
- MntA mediates the consecutive transfer of three AMPs to a tyrosine residue near HepT's RNase domain.
- The conserved GSX10DXD motif in MntA is essential for polyadenylylation and HepT neutralization.
- Polyadenylylation by MntA is crucial for reducing HepT's RNase toxicity.
Conclusions:
- HepT/MntA represents a novel type of toxin/antitoxin system.
- Polyadenylylation-dependent neutralization is a prevalent mechanism in bacterial and archaeal toxin/antitoxin systems.
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