Molecular Cloning and Characterization of a Human Multisubstrate Specific Nucleotide-sugar Transporter Homologous to Drosophila fringe connection

Abstract

Nucleotide-sugar transporters are crucial components in the synthesis of glycoconjugates. We identified a novel human nucleotide-sugar transporter gene, hfrc1, which is homologous to Drosophila melanogaster fringe connection, Caenorhabditis elegans sqv-7, and human UGTrel7. HFRC1 was localized within the Golgi apparatus following its transient expression in HCT116 cells. In human tissues, hfrc1 and UGTrel7 exhibited similar tissue distributions, although hfrc1 transcripts showed a 10 times greater abundance than those of UGTrel7. The heterologous expression of HFRC1 in the yeast revealed the multisubstrate specific transport activity of HFRC1 (for UDP-N-acetylglucosamine (UDP-GlcNAc), UDP-glucose (UDP-Glc), and GDP-mannose (GDP-Man), with apparent K m values of 8.0, 2.1, and 0.14 M, respectively). In the mammalian cells, HFRC1 transported UDP-GlcNAc and UDP-Glc, but not GDP-Man. Overexpression of the hfrc1 gene in HCT116 cells modulated the cell surface heparan sulfate expression status. These results suggest that HFRC1 takes part in the synthesis of heparan sulfate by regulating the level of UDP-GlcNAc, a donor substrate for the heparan sulfate synthases.

The Drosophila Neurally Altered Carbohydrate Mutant Has a Defective Golgi GDP-fucose Transporter

“…The residual levels of monofucosylated N-glycans indicate that nac 1 flies are still able to transport some GDP-fucose into the Golgi. This suggests the presence of an alternative, functionally redundant GDP-fucose transport mechanism, a notion that is supported by the results of another study, in which faint anti-HRP and A. aurantia lec- (69), and humans (human FRC1 and hUGTrel7) (68,70).…”

Role of UDP-N-Acetylglucosamine (GlcNAc) and O-GlcNAcylation of Hyaluronan Synthase 2 in the Control of Chondroitin Sulfate and Hyaluronan Synthesis

“…GAGosome) (42,43), and it was demonstrated that overexpression of the nucleotide-sugar transporter HFRC1 induced accumulation of heparan sulfate (44). Interestingly, UDP-glucose pyrophosphorylase, which produces UDPglucose from glucose 1-phosphate, was reported to be O-GlcNAcylated (45).…”

Independent and simultaneous translocation of two substrates by a nucleotide sugar transporter

“…Since then, multisubstrate transporters of nucleotide sugars have been described in several organisms, including L. donovani (29), D. melanogaster (12), humans (20), E. histolytica (23), etc. A competitive substrate translocation mechanism has been since then assumed but not proven in any other study.…”

“…Although both transporters transport multiple substrates, it is only for SQV-7, the first described multisubstrate nucleotide sugar transporter, that competition among these substrates has been demonstrated. In all other cases, including mammals (20,21), insects (11,12), plants (22), and protista (23), where multiple substrates have been described for a given nucleotide sugar transporter, a competitive transport mechanism has been assumed but not shown.…”