A Golgi oxygen sensor controls intestinal mucin glycosylation

Plain language summary

Intestinal mucin glycosylation is important for mucus‐bacterial homeostasis and is altered in disease. In this issue of The EMBO Journal, Ilani et al (2022) identify the Golgi enzyme quiescin sulfhydryl oxidase 1 (QSOX1) as a novel mucus regulator by controlling mucin sialylation.

The Golgi enzyme quiescin sulfhydryl oxidase 1 (QSOX1) emerges as a novel mucus regulator by controlling mucin sialylation.

The gel‐forming mucins constitute the skeleton of mucus, formed by large disulfide‐bonded polymers that are highly glycosylated. MUC2, the main intestinal mucin, forms two‐layered mucus in the colon. The inner mucus layer acts as an impenetrable barrier to bacteria in healthy individuals (Johansson et al, 2008). The outer mucus layer harbors the commensal bacteria that utilize the mucin glycans as their energy supply. The presence of a healthy anaerobic bacterial community in this outer mucus layer is important for the preservation of health.

Mucins and the blood clotting protein von Willebrand factor (VWF) are structurally closely related and both form a disulfide‐bonded dimer in their far C‐terminal cysteine‐knot domain during biosynthesis in the endoplasmic reticulum. As the secretory pathway cannot handle large polymers, the N‐terminal disulfide bond polymerization has to be delayed until the mucins or VWF have passed into the late Golgi or trans‐Golgi network (TGN). The mechanisms mediating mucin polymerization at the Golgi have remained poorly understood. Ilani et al (2022) hypothesized that the Golgi‐resident enzyme quiescin sulfhydryl oxidase 1 (QSOX1), which is especially abundant in intestinal goblet cells (Nystrom et al, 2021), is responsible for this oligomerization. Due to the insoluble nature of the MUC2 mucin polymers, Ilani et al (2022) instead analyzed whether either the MUC5B mucin or VWF was able to form their classical polymeric ladders in mice lacking QSOX1. Both showed identical ladders in wild‐type and knockout animals, thus disproving their hypothesis and indicating that further studies are required to uncover the mechanisms mediating mucin polymerization in the Golgi.

The Golgi apparatus houses the cellular glycosylation machinery, where the glycosyltransferases are geographically arranged according to the order of monosaccharide building block addition (Rabouille et al, 1995). The localization of the glycosyltransferases in the Golgi stack depends on its decreasing pH gradient, elimination of which alters the glycosyltransferase localization that causes simplified glycan structures (Axelsson et al, 2001). In a recent seminal publication, Kellokumpu and coworkers (Hassinen et al, 2019) found that hypoxia inhibited terminal sialylation at the Golgi. Interestingly, they found that hypoxia was accompanied with the loss of two surface disulfide bonds in the terminal sialylating enzyme α‐2,6‐sialyltransferase (ST6Gal1), rendering the enzyme inactive. Hassinen et al hypothesized that QSOX1 could be the enzyme responsible for disulfide bond formation in ST6Gal1. Ilani et al (2022) have now taken up the open question and shown that the mice lacking QSOX1 indeed show altered glycosylation patterns. Using lectin staining, the group demonstrated that mucins in QSOX1 knockout mice show decreased sialylation. Furthermore, they show that QSOX1 is required for the formation of the activity‐regulating disulfide bonds in ST6Gal1 and a subset of other Golgi glycosyltransferases (Ilani et al, 2022).

QSOX1 uses molecular oxygen to oxidize two sulfhydryl groups to generate a disulfide bond and hydrogen peroxide (Fig 1A). This suggests that QSOX1 can act as a sensor of oxygen tension and the redox conditions in the goblet cell and by this regulate specific glycosyltransferase activities. As the biosynthesis of glycans in the Golgi is sequential, activity of individual glycosyltransferases can affect levels of many glycans. The observation of QSOX1 regulation of glycosylation brings into focus the poorly understood regulation of glycosylation and shows that analyzing glycosylation enzyme mRNA expression alone is insufficient.

Figure 1. Enzymatic reaction catalyzed by QSOX1 and its effect on mucin glycosylation in the Golgi apparatus of intestinal goblet cells.

(A) QSOX1 catalyzes disulfide bond formation in Golgi sialyltransferases. (B) Left: Golgi‐localized sialyltransferase ST6GAL1 is inactive when the disulfide bonds Cys142‐Cys406 and Cys353‐Cys364 are reduced. Right: disulfide bond formation at these residues activates ST6GAL1, leading to sialylation of the intestinal mucin MUC2 in the goblet cells.

Why should mucin glycosylation be controlled or influenced by oxygen tension? A possible clue to this might be found in the large intestine, where the glycan repertoire is very important for the homeostatic balance between bacteria and the host. The luminal bacteria of the colon are more or less obligatory anaerobic, a milieu that is important to maintain and control also by the host. However, we do not know if and how the host might record and control the bacterial composition to make sure it maintains an anaerobic environment. Mechanisms for host selection of normal commensal bacteria are also poorly understood, although early experiments with bacterial transfer between different species strongly show important host effects (Rawls et al, 2006). The exposed surface environment of the gut is almost exclusively glycan‐rich and varies between species, thus arguing for glycans playing a major role in the selection process. This is likely accomplished by the variable capacity of different bacteria to degrade and utilize the glycans (Luis et al, 2021). However, even more important is likely the capacity of bacteria to attach to the surface‐associated inner mucus layer and in this way avoid removal by peristalsis. Molecular details for this are not yet understood but will likely include binding to complex glycan surfaces made up by glycosylated mucin domains. The outer rim of the inner colon mucus layer is accessible by bacteria due to the expanded nature of the outer mucus layer and will retain glycans as biosynthesized by the host before the bacterial hydrolases have altered their surface. In line with this discussion, Ilani et al (2022) show that the mice lacking QSOX1 had not only altered glycosylation, but also an altered microbiota (Ilani et al, 2022).

Glycan analyses of colonic mucins reveal less sialylation and more sulfation of the MUC2 glycans toward the distal end of the large intestine in both mouse and humans (Robbe et al, 2003; Holmen‐Larsson et al, 2013). Interestingly, this is not reflected in the levels of glycosyltransferases, as both sialyl‐ and sulfotransferase expression increases distally (van der Post & Hansson, 2014). This raises the possibility that the anaerobic colon lumen and its sensing by QSOX1 could modulate transferase activities and lead to decreased levels of sialylation.

The observations by Kellokumpu et al and now further by Ilani et al suggest an interesting model of how glycosylation might be controlled by oxygen tension and offer a glimpse into possible molecular mechanisms. Substantial future work on both host and bacteria will be required to more fully understand the physiological importance of QSOX1 in health and disease.

The EMBO Journal (2023) 42: e113013