Loss of srf-3-encoded Nucleotide Sugar Transporter Activity in Caenorhabditis elegans Alters Surface Antigenicity and Prevents Bacterial Adherence

Abstract

During the establishment of a bacterial infection, the surface molecules of the host organism are of particular importance, since they mediate the first contact with the pathogen. In Caenorhabditis elegans, mutations in the srf-3 locus confer resistance to infection by Microbacterium nematophilum, and they also prevent biofilm formation by Yersinia pseudotuberculosis, a close relative of the bubonic plague agent Yersinia pestis. We cloned srf-3 and found that it encodes a multitransmembrane hydrophobic protein resembling nucleotide sugar transporters of the Golgi apparatus membrane. srf-3 is exclusively expressed in secretory cells, consistent with its proposed function in cuticle/surface modification. We demonstrate that SRF-3 can function as a nucleotide sugar transporter in heterologous in vitro and in vivo systems. UDP-galactose and UDP-N-acetylglucosamine are substrates for SRF-3. We propose that the inability of Yersinia biofilms and M. nematophilum to adhere to the nematode cuticle is due to an altered glycoconjugate surface composition of the srf-3 mutant.

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

“…Transport of UDP-GlcNAc and UDP-GalNAc were saturable with apparent K m values of 25.2 and 24.9 M, respectively. These results are in close agreement with previously described apparent K m values for other nucleotide sugar transporters (8).…”

“…This facilitates assays of nucleotide sugar transport when a putative nucleotide sugar transporter from a different species is expressed in S. cerevisiae. This approach has been used by us and others to determine substrate specificity of transporters from mammals (26), C. elegans (7,8), plants (22), and protists (23). As shown in Fig.…”

“…Mutants of nucleotide sugar transporters have developmental phenotypes and have been described in Caenorhabditis elegans (7,8), yeast (9), Leishmania donovani (10), Drosophila melanogaster (11,12), cattle (13), and humans (14)(15)(16). Mutations in the transporters of UDP-N-acetylglucosamine (UDP-GlcNAc) in Holstein cows and GDP-fucose in humans give rise to complex vertebral malformation and leukocyte adhesion deficiency syndrome II, respectively.…”

“…Although mutants deficient in SRF-3 activity do not show morphological phenotypes when grown on E. coli, they are resistant to infection by the bacterial pathogens Microbacterium nematophilum and Yersinia pseudotuberculosis, whereas the wildtype nematodes are not. SRF-3 has a narrow tissue expression pattern, and mutants have a decrease in O-and N-linked galactose-containing glycoconjugates (8,19).…”

Yersinia pestis Biofilm in the Flea Vector and Its Role in the Transmission of Plague

“…For example, C. elegans mutants with an altered surface cuticle do not accumulate Yersinia biofilm (Höflich et al 2004;Joshua et al 2003). Thus, initiation and development of the biofilm growth phenotype can be multifactorial and environment-specific.…”