An intra-bacterial activity for a T3SS effector

Abstract

Many Gram-negative bacterial pathogens interact with mammalian cells by using type iii secretion systems (T3SS) to inject virulence proteins into host cells. A subset of these injected protein 'effectors' are enzymes that inhibit the function of host proteins by catalyzing the addition of unusual posttranslational modifications. The E. coli and Citrobacter rodentium NleB effectors, as well as the Salmonella enterica SseK effectors are glycosyltransferases that modify host protein substrates with N-acetyl glucosamine (GlcNAc) on arginine residues. This post-translational modification disrupts the normal functioning of host immune response proteins. T3SS effectors are thought to be inactive within the bacterium and fold into their active conformations after they are injected, due to the activity of chaperones that keep the effectors in a structural state permissive for secretion. While performing mass spectrometry experiments to identify glycosylation substrates of nleB orthologs, we unexpectedly observed that the bacterial glutathione synthetase (GshB) is glycosylated by NleB on arginine residue R256. NleB-mediated glycosylation of GshB resulted in enhanced GshB activity, leading to an increase in glutathione production, and promoted C. rodentium survival in oxidative stress conditions. these data represent, to our knowledge, the first intra-bacterial activity for a T3SS effector and show that arginine-GlcnAcylation, once thought to be restricted to host cell compartments, also plays an important role in regulating bacterial physiology.

Tubulin Folding Cofactor TBCB is a Target of the Salmonella Effector Protein SseK1

“…In addition, NleB/SseK proteins may catalyze the glycosylation of endogenous bacterial proteins. A recent report has shown that NleB1 from enterohemorrhagic E. coli, NleB from C. rodentium, and SseK1 from S. enterica, can glycosylate the bacterial protein GshB (glutathione synthetase) [31]. This glycosylation enhances GshB-mediated production of glutathione, which provides resistance to oxidative stress, suggesting a new role for these effectors in promoting the survival of bacteria under these conditions.…”

Type III Secretion Effectors with Arginine N-Glycosyltransferase Activity

“…Interestingly, a recent report has shown that NleB1 can also glycosylate a bacterial protein, GshB (glutathione synthetase) on Arg 256 . In fact, this was the most abundant arginine-GlcNAcylated target enriched from samples of HEK293T cells infected with EHEC [179]. GshB was also glycosylated in vitro and in vivo by C. rodentium NleB and in vitro by Salmonella SseK1.…”

Bacteria-Catalyzed Arginine Glycosylation in Pathogens and Host

“…Remarkably, in more recent work, Wuhan Xiao and co-workers discovered that HIF-1α was GlcNAcylated at a conserved arginine (Arg18) during C. rodentium or EPEC infection, and the modification enhanced HIF-1α transcriptional activity, thus inducing downstream glucose metabolism-associated gene [such as glucose transporter 1 (GLUT1) gene] expression to alter host glucose metabolism (Xu et al, 2018). Additionally, a more recently study has shown that bacterial glutathione synthetase (GshB) was GlcNAcylated by C. rodentium NleB on Arg256 (El Qaidi et al, 2020). Further, NleBmediated GlcNAcylation of GshB contributed to C. rodentium survival in oxidative stress conditions (El Qaidi et al, 2020).…”