Molecular Cloning and Characterization of the Caenorhabditis elegans α1,3-Fucosyltransferase Family

The genome of Caenorhabditis elegans encodes five genes with homology to known α1,3 fucosyltransferases (α1,3FTs), but their expression and functions are poorly understood. Here we report the molecular cloning and characterization of these C. elegans α1,3FTs (CEFT-1 through -5). The open-reading frame for each enzyme predicts a type II transmembrane protein and multiple potential N-glycosylation sites. We prepared recombinant epitope-tagged forms of each CEFT and found that they had unusual acceptor specificity, cation requirements, and temperature sensitivity. CEFT-1 acted on the N-glycan pentasaccharide core acceptor to generate Manα1-3(Manα1-6)Manβ1-4GlcNAcβ1-4(Fucα1-3)GlcNAcβ1-Asn. In contrast, CEFT-2 did not act on the pentasaccharide acceptor, but instead utilized a LacdiNAc acceptor to generate GalNAcβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, which is a novel activity. CEFT-3 utilized a LacNAc acceptor to generate Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc without requiring cations. CEFT-4 was similar to CEFT-3, but its activity was enhanced by some divalent cations. Recombinant CEFT-5 was well expressed, but did not act on available acceptors. Each CEFT was optimally active at room temperature and rapidly lost activity at 37 °C. Promoter analysis showed that CEFT-1 is expressed in C. elegans eggs and adults, but its expression was restricted to a few neuronal cells at the head and tail. We prepared deletion mutants for each enzyme for phenotypic analysis. While loss of CEFT-1 correlated with loss of pentasaccharide core activity and core α1,3-fucosylated glycans in worms, loss of other enzymes did not correlate with any phenotypic changes. These results suggest that each of the α1,3FTs in C. elegans has unique specificity and expression patterns.