The Caenorhabditis elegans gene, gly-2, can rescue the N-acetylglucosaminyltransferase V mutation of Lec4 cells.
UDP-N-acetylglucosamine:alpha-6-d-mannoside beta-1,6-N-acetylglucosaminyltransferase V (GlcNAc-TV) is a regulator of polylactosamine-containing N-glycans and is causally involved in T cell regulation and tumor metastasis. The Caenorhabditis elegans genome contains a single orthologous gene, gly-2, that is transcribed and encodes a 669-residue type II membrane protein that is 36.7% identical to mammalian GlcNAc-TV (Mgat-5). Recombinant GLY-2 possessed GlcNAc-TV activity when assayed in vitro, and protein truncations demonstrated that the N-terminal boundary of the catalytic domain is Ile-138. gly-2 complemented the Phaseolus vulgaris leucoagglutinin binding defect of Chinese hamster ovary Lec4 cells, whereas GLY-2(L116R), an equivalent mutation to that which causes the Lec4A phenotype, could not. We conclude that the worm gene is functionally interchangeable with the mammalian form. GlcNAc-TV activity was detected in wild-type animals but not those homozygous for a deletion allele of gly-2. Activity was restored in mutant animals by an extrachromosomal array that encompassed the gly-2 gene. Green fluorescent protein reporter transgenes driven by the gly-2 promoter were expressed by developing embryos from the late comma stage onward, present in a complex subset of neurons in larvae and, in addition, the spermathecal and pharyngeal-intestinal valves and certain vulval cells of adults. However, no overt phenotypes were observed in animals homozygous for deletion alleles of gly-2.
Warren
Charles E
CE
Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario M5G 1X5, Canada.
Krizus
Aldis
A
Roy
Peter J
PJ
Culotti
Joseph G
JG
Dennis
James W
JW
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D016466:CHO Cells; D017173:Caenorhabditis elegans / Q000235:genetics*; D029742:Caenorhabditis elegans Proteins / Q000096:biosynthesis / Q000235:genetics*; D002874:Chromosome Mapping; D006224:Cricetinae; D018076:DNA, Complementary / Q000378:metabolism; D017353:Gene Deletion; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D016681:Genome, Fungal; D049452:Green Fluorescent Proteins; D006720:Homozygote; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008861:Microsomes / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism*; D010641:Phenotype; D011061:Poly A / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D019076:Transgenes
D029742:Caenorhabditis elegans Proteins; D018076:DNA, Complementary; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C459808:gly-2 protein, C elegans; D049452:Green Fluorescent Proteins; D011061:Poly A; D017351:N-Acetylglucosaminyltransferases; C052254:alpha-1,6-mannosylglycoprotein beta 1,6-N-acetylglucosaminyltransferase