Fucosylation in the gut. The secret(or) of blood sugars

Fucosylation is a type of glycosylation involving an enzymatic addition of fucose residues to a molecule. Fucosyltransferases transfer fucose from GDP-Fuc to Gal in an α(1,2)-linkage and to GlcNAc in α(1,3)-, α(1,4)-, or α(1,6)-α(1,2)-linkages (99). In humans, the FUT gene family comprises 13 different genes encoding fucosyltransferases displaying a complex tissue- and cell type–specific pattern of an expression (100). Alpha-2-fucosyltransferases (encoded in mammals by Fut1, Fut2, and Sec1 genes) are enzymes required for the biosynthesis of the terminal glycan motif Fucα2-Galβ-R (see Table 1, Figure 8B-C). Human FUT1 (H) and FUT2 (Secretor) genes are responsible for synthesis of ABH and Lewis histo-blood groups (Figure 6).

Upon glycosylation, α(1,2)-fucosylated proteins and lipids (101) can be directly secreted into the gut lumen or can be anchored to the apical side of the mucosal epithelium (102). Indeed, polyfucosylated (mostly Fucα1,2) N-linked tri- and tetra-antennary glycopeptides with blood group determinants were found in abundance in human small intestinal epithelial cells (103). In individuals with the functional α-2-fucosyltransferase FUT2 (so-called “secretor” phenotype, Se), ABH blood group antigens (analogous to those found on a surface of erythrocytes) are expressed on epithelial cells of higher respiratory tract, lower genito-urinary tract, on gastrointestinal epithelia and on mucins (Table 1) (104). For example, “secretors” produce Lewis^b and H type 1 antigens (mirroring blood group O); H antigen structure can be further decorated by another glycosyltransferases with GalNAc- or Gal-residues thus creating A or B antigens (Figure 6A-B).

In contrast, “non-secretors” (Se⁰) subjects are homozygous for the FUT2-inactivating nonsense mutation and, thus, express only the Lewis^a antigen (with FUT3-dependent

19 Fucα1,4) instead of Lewis^b and ABH antigens (105) (Figure 6A-B). Moreover, the “weak-secretor” phenotype (Se^w) which is characterized by an expression of both Lewis^a and Lewis^b antigens (Figure 6C-D) has been identified in Taiwanese, Polynesians, Japanese, and Australian aborigines (106). Se^w phenotype is the result of a weak (mutated) form of the α-2-fucosyltransferase (107). To conclude, a secretor status is determined by a concurrent action of two different enzymes, Secretor (FUT2) and Lewis (FUT3). Of note, Serpa et al. reported that only nine out of 47 Lewis-negative (i.e., FUT3-negative) individuals lacked the expression of Lewis antigens in gastric mucosa (Le^a-, Le^b-) indicating a possible contribution of another α-1,3/4 fucosyltransferase to Lewis antigens biosynthesis in the mucosa (108).

Naturally occurring polymorphisms of FUT1 and FUT2 genes (due to fully or partially inactivating mutations) have significant epidemiological and functional consequences, highlighting the importance of α(1,2)-fucosylated glycans. Lack of both functional FUT1 and FUT2 alleles results in the very rare Bombay blood group (109) characterized by an absence of ABH antigens on erythrocytes (regardless of the blood group genotype) and in secretions (110). Individuals with the Para-Bombay phenotype are also FUT1-negative, but do express the FUT2–encoded α(1,2)-fucosyltransferase, and thus “secretors” (111).

Figure 6. Fucosylated blood group antigens and associated secretor phenotypes. (A) The α(1.2)-fucosylated (in red) H type 1 and Lewis^b antigens correspond to blood group O. Group A and B antigens have additional GalNAc or Gal residues (blue), respectively. Sialylated Lewis antigens (depicted in pink) are up-regulated during inflammation and infection. (B) Biosynthesis pathways for blood group antigens with corresponding Se phenotypes. (C) Humans of Se, Se⁰, or Se^w phenotype are characterized by the presence/absense of ABH and Lewis^a antigens in salivary, milk, and GI tract secretions. (D) An activity of α1.2fucosyltransferase defines the secretor status. Reproduced from (112) under CC0 1.0 Universal Public Domain Dedication (no copyright).

20 FUT2 gene polymorphism is more common: it is estimated that approximately one-fifth of the global population are “non-secretors” unable to express α(1,2)-fucosylated glycans on mucosa (102). FUT2 gene, located on Chr19q13.33, has length of 9,980 bp, and it is composed of two exons separated by a 6,865-bp intron. Two functional alleles of FUT2 are commonly found in many populations: one is a so‐called reference allele (Se), and the other is the Se³⁵⁷ with a 357 C→T synonymous SNP (113). Nevertheless, more than 20 different single nucleotide polymorphisms (SNPs) in FUT2 conferring the “non-secretor” phenotype were described (114).

Similarly to the population-specific Se^w phenotype, the frequency of Se⁰ varies in different ethnic groups. For example, in populations of Caucasian, African, and central Asian descent, the most common nonsense SNP is a G→A substitution at base pair 428 (rs601338) (105). In contrast, Se³⁸⁵, an A→T substitution at base pair 385 (rs1047781), is the most frequent cause of the “non-secretor” phenotype in east Asians (107). Notably, within their respective populations, both aforementioned SNPs are distributed at similar frequencies (≈20%) (113). It seems that, at least in Eurasian and African populations, many FUT2 variants had a long history of (probably pathogen-driven) balancing selection ensuring both Se and Se⁰ had an advantage (115). It is estimated that the Se⁴²⁸ mutation is at least 1.87 million years old (116). Fast spreading of the most recent, H8 haplotype (Se³⁸⁵), which appeared in East Asia about 256,000 years ago, might reflect a positive selection (113). To conclude, the world-wide distribution of Se⁰ phenotypes may be a result of on-going selective pressure providing an advantage for “non-secretor” in the certain cases.

Possible associations between the secretor status and susceptibility to infections and diseases have been extensively studied. For instance, a recent study claimed that FUT2 SNPs rs601338 (Se⁴²⁸) and rs602662 were associated with a higher risk of diarrhea and respiratory illnesses in infants, while a longer period of breastfeeding reduced risk of diarrhea, independent of FUT2 genotype (117). Furthermore, “secretors” were shown to be more susceptible to norovirus, rotavirus, HIV, Helicobacter pylori, and graft-versus-host disease, while “non-secretors” are more prone to Candida albicans, Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenza infections, as well as to gastric cancer, primary sclerosing cholangitis, and Crohn’s disease (reviewed in 102).

Some of the pathogens mentioned above can directly bind and adhere to α(1,2)-fucose residues (discussed in the Chapter 1.4.2), which explains the observed differences in colonization. Interestingly, while Fut2 gene expression affected the gut community assembly

21 in mice (118), no strong correlation between a secretor status and the composition of the human gut microbiota was detected in a twin cohort study (119). Finally, it should be noted, that “non-secretors” had increased levels of sialylated glycans (incl. Sialyl-Lewis^x) expressed on MUC5B isolated from saliva, probably, due to reduced competition between glycosyltransferases (120). Enhanced sialylation may mask or aggravate Se⁰-dependent phenotypes.