Inhibition of O‐GlcNAcylation protects from Shiga toxin‐mediated cell injury and lethality in host

Abstract

Shiga toxins (Stxs) produced by enterohemorrhagic Escherichia coli (EHEC) are the major virulence factors responsible for hemorrhagic colitis, which can lead to life-threatening systemic complications including acute renal failure (hemolytic uremic syndrome) and neuropathy. Here, we report that O-GlcNAcylation, a type of posttranslational modification, was acutely increased upon induction of endoplasmic reticulum (ER) stress in host cells by Stxs. Suppression of the abnormal Stx-mediated increase in O-GlcNAcylation effectively inhibited apoptotic and inflammatory responses in Stx-susceptible cells. The protective effect of O-GlcNAc inhibition for Stx-mediated pathogenic responses was also verified using three-dimensional (3D)cultured spheroids or organoids mimicking the human kidney. Treatment with an O-GlcNAcylation inhibitor remarkably improved the major disease symptoms and survival rate for mice intraperitoneally injected with a lethal dose of Stx. In conclusion, this study elucidates O-GlcNAcylation-dependent pathogenic mechanisms of Stxs and demonstrates that inhibition of aberrant O-GlcNAcylation is a potential approach to treat Stx-mediated diseases.

Intestinal mucus-derived metabolites modulate virulence of a clade 8 enterohemorrhagic Escherichia coli O157:H7

“…First, the colonic epithelium presents high levels of O-NAG-Acylations on the cell surface of enterocytes, which could be sensed by E. coli O157:H7 and represent an indicator of proximity to the epithelium and, therefore, of a proinflammatory environment. Furthermore, Stx2 is known to induce overexpression of O-NAG-Acylations in host cell proteins, conferring a boost to this signal ( Lee et al., 2022 ). Another possible explanation for the increase in Stx2 in the presence of NAG may be associated to the evolutionary origin of the incorporation of the Stx2-associated phage in E. coli , which is accepted to be related to the toxicity that it produces on its natural predators and competitors, such as protists and fungi ( Steinberg and Levin, 2007 ; Koudelka et al., 2018 ).Taking into account that chitin is the main component of the fungal cell wall ( Min et al., 2020 ), that this molecule is a polymer of NAG easily catabolizable to its constituent monosaccharides ( Hui et al., 2020 ), and that NAG is already used as a modulator of virulence by E. coli O157:H7 ( Sohanpal et al., 2004 ; Konopka, 2012 ; Le Bihan et al., 2017 ; Sicard et al., 2018 ) we hypothesize that a high amount of this sugar could be used by E. coli O157:H7 as a signal indicating an unfavorable environment related to a prevalence of fungi in the intestinal environment.…”

Host Cellular Response during Enterohaemorrhagic Escherichia coli Shiga Toxin Exposure

“…Consequently, the expression levels of CHOP, XBP1s, and death receptor 5 (DR5) were increased by type 1 Shiga toxin stimulation, leading to activation of caspase 3 and 8. Shiga toxin type 2 activates the UPR response, which may induce CHOP and DR5 expression, leading to apoptosis [3,102]. Interestingly, the individual A and B subunits of Shiga toxin and a catalytically deficient toxin (Stx mut ) cannot induce UPR responses, suggesting a key role for the enzymatic activity of the A subunit [101].…”

Shiga toxin (Stx) type 2‐induced increase in O‐linked N‐acetyl glucosamine protein modification: a new therapeutic target?

“…An anticomplement therapy that is successful in noninfectious HUS, Eculizumab, has not consistently demonstrated similar benefits for STEC‐related HUS (Mühlen & Dersch, 2020). However, Lee et al describe a novel strategy for interrupting Stx2a‐based damage and death in multiple cell types and organoids, and demonstrated protection of mice injected with Stx2a by an inhibitor of O‐linked N‐acetyl glucosamine protein modification O‐GlcNAcylation, OSMI‐1 (Lee et al , 2021). Protein modification by the addition of O‐GlcNAc affects many cellular processes, including the cell cycle and stress responses (Martinez et al , 2017; Estevez et al , 2020).…”

“…The decision to test whether OSMI‐1 could protect from Stx2a‐mediated damage came from the authors’ original finding that THP‐1 cells treated with Stx2a had increased levels of O ‐ GlcNAcylation associated with protein in cell lysates (Lee et al , 2021). This result revealed yet another cellular process altered by Stx intoxication.…”

“…The increase in O‐GlcNAcylation observed by Lee et al may well be a reaction by the cell to ER stress induced by Stx2a; as such, O‐GlcNAcylation has been shown to decrease injury due to ER stress [see review (Martinez et al , 2017)]. Importantly, the authors showed that a catalytically inactive version of Stx2a that cannot target the ribosome was unable to increase O‐GlcNAcylation (Lee et al , 2021). This study demonstrated that in addition to an ER stress response, that an overall increase in O‐GlcNAc levels occurred, and that both pro‐inflammatory (p65) and pro‐apoptotic (Akt and Bad) signaling proteins were O‐GlcNAcylated, their phosphorylation status altered, and function increased.…”