Methionine oxidation activates a transcription factor in response to oxidative stress

Abstract

Oxidant-mediated antibacterial response systems are broadly used to control bacterial proliferation. Hypochlorite (HOCl) is an important component of the innate immune system produced in neutrophils and specific epithelia. Its antimicrobial activity is due to damaging cellular macromolecules. Little is known about how bacteria escape HOCl-inflicted damage. Recently, the transcription factor YjiE was identified that specifically protects Escherichia coli from HOCl killing. According to its function, YjiE is now renamed HypT (hypochlorite-responsive transcription factor). Here we unravel that HypT is activated by methionine oxidation to methionine sulfoxide. Interestingly, so far only inactivation of cellular proteins by methionine oxidation has been reported. Mutational analysis revealed three methionines that are essential to confer HOCl resistance. Their simultaneous substitution by glutamine, mimicking the methionine sulfoxide state, increased the viability of E. coli cells upon HOCl stress. Triple glutamine substitution generates a constitutively active HypT that regulates target genes independently of HOCl stress and permanently down-regulates intracellular iron levels. Inactivation of HypT depends on the methionine sulfoxide reductases A/B. Thus, microbial protection mechanisms have evolved along the evolution of antimicrobial control systems, allowing bacteria to survive within the host environment.

Selenoproteins: Molecular Pathways and Physiological Roles

“…Similar to other reversible posttranslational modifications, e.g., phosphorylation of tyrosine residues in proteins, methionine oxidation and reduction were proposed to play regulatory roles (144,220). For example, methionine oxidation has been shown to activate the transcription factor, HypT, which is involved in the bacterial response to hypochlorite (88). Moreover, oxidation of methionine residues has been shown to activate calcium/calmodulin (Ca 2ϩ /CaM)-dependent protein kinase II (CaMKII) in the absence of Ca 2ϩ /CaM, whereas MsrA activity could reverse this effect (93).…”

Convergent signaling pathways—interaction between methionine oxidation and serine/threonine/tyrosine O-phosphorylation

“…However, recently, an instance was reported where Met oxidation led to the activation of a protein function. The oxidation of multiple Met residues in the hypochlorite-responsive transcription factor (HypT) was found to stimulate the role that it plays in preventing hypochlorite-induced stress in Escherichia coli (Drazic et al 2013). Based upon the results from structural modeling of HypT, and comparison with the redox active and structurally similar E. coli transcription factor OxyR, the authors speculated that oxidation-mediated conformational rearrangements in HypT alter contacts within the dimer/ tetramer interface and facilitate binding to target DNA.…”

“…In most instances, Met oxidation has a negative effect on subsequent O-phosphorylation (Table 1), although the ultimate downstream readout can be positive. However, as is the case of HypT (Drazic et al 2013), there are some events immediately downstream where Met oxidation has an immediate positive regulatory effect (e.g., Met oxidation allows Ca 2+ /CaM-independent activation of CaMKII; Erickson et al 2008).…”

Tetramers Are the Activation-competent Species of the HOCl-specific Transcription Factor HypT

“…Whereas Hsp33, NemR, and HypR are activated by oxidation of cysteine thiols (7,8,11), HypT is activated by HOCl by simultaneous oxidation of three methionines (Met-123, Met-206, and Met-230) to methionine sulfoxide, thus forming the active HypT species (10). This active state can be mimicked by methionine-to-glutamine substitution, leading to the constitutively active mutant HypT M123,206,230Q (10). Both active HypT and the constitutively active mutant show strong DNA binding.…”

“…However, it dissociates into dimers and tetramers upon incubation with DNA in vitro, thus forming the DNA-binding species (9). Of note, given the reduced state of the protein, DNA binding is only weak (9,10). This DNA-induced dissociation is independent of the DNA used and occurs with nonspecific DNA and target DNA alike (9).…”