The Methanosarcina acetivorans thioredoxin system activates DNA binding of the redox-sensitive transcriptional regulator MsvR

Abstract

The production of biogas (methane) by anaerobic digestion is an important facet to renewable energy, but is subject to instability due to the sensitivity of strictly anaerobic methanogenic archaea (methanogens) to environmental perturbations, such as oxygen. An understanding of the oxidant-sensing mechanisms used by methanogens may lead to the development of more oxidant tolerant (i.e. stable) methanogen strains. MsvR is a redox-sensitive transcriptional regulator that is found exclusively in methanogens. We show here that oxidation of MsvR from Methanosarcina acetivorans (MaMsvR) with hydrogen peroxide oxidizes cysteine thiols, which inactivates MaMsvR binding to its own promoter (PmsvR). Incubation of oxidized MaMsvR with the M. acetivorans thioredoxin system (NADPH, MaTrxR, and MaTrx7) results in reduction of the cysteines back to thiols and activation of PmsvR binding. These data confirm that cysteines are critical for the thiol-disulfide regulation of PmsvR binding by MaMsvR and support a role for the M. acetivorans thioredoxin system in the in vivo activation of MaMsvR. The results support the feasibility of using MaMsvR and PmsvR, along with the Methanosarcina genetic system, to design methanogen strains with oxidant-regulated gene expression systems, which may aid in stabilizing anaerobic digestion.

A Novel F420-dependent Thioredoxin Reductase Gated by Low Potential FAD

“…However, it has also been investigated significantly in the anaerobic organisms (6 -24) focusing on both the characteristics of the Trxs and TrxRs (6,8,9,(11)(12)(13)(15)(16)(17)(18)(19)(20)(21)(22)(23) and their physiological roles (6,7,10,14,20). For example, the isolation of the putative Trx targets via Trxaffinity chromatography and the results from activation assays with select purified and deactivated enzymes have established that in Chlorobaculum tepidum, a green sulfur bacterium and an anaerobic phototroph, the thioredoxin system implements post-translational control on the tricarboxylic acid or the TCA cycle and sulfur metabolism and assists in the defense against oxidative stress (7).…”

“…Recently, a proteomics study has shown that there is clear potential for a Trx-based global redox regulation of metabolism, including methanogenesis, in Methanocaldococcus jannaschii, a strictly anaerobic and deeply rooted hyperthermophilic methanogenic archaeon that lives in deep-sea hydrothermal vents (20). Two reports also describe the activation of deactivated methanogen proteins by thioredoxin (10,20).…”

Redox and Thiols in Archaea

“…In M. acetivorans, treatment of MsvR with H 2 O 2 results in oxidation of cysteine thiols, preventing binding of MsvR to promoters. Incubation of oxidized MsvR with the M. acetivorans TRX system, consisting of NADPH, TRX reductase and one of the 7 TRXs, leads to reduction of the cysteines and binding to its own promoter [111].…”

Thiol‐based redox sensing in the methyltransferase associated sensor kinase RdmS in Methanosarcina acetivorans

“…M. acetivorans was shown to possess a single thioredoxin reductase TrxR and seven Trx homologues (Trx1-7), at least three of which are functional (McCarver and Lessner, 2014). In this regard, it has previously been shown that the Trx system is able to reduce disulfides back to thiols in the redox-sensitive transcriptional regulator MsvR of M. acetivorans (Sheehan et al, 2015).…”