UDP-GlcNAc: alpha-3-D-mannoside beta1,2-N-acetylglucosaminyltransferase I (GnT I) is a Golgi-resident enzyme which transfers a GlcNAc residue in beta1,2 linkage to the Manalpha1,3Manbeta-terminus of (Manalpha1,6(Manalpha1,3)Manalpha1,6)(Manalpha1,3)Manbeta1, 4GlcNAcbeta1,4G lcNAc-Asn-protein, thereby initiating the synthesis of hybrid N-glycans. Three Caenorhabditis elegans genes homologous to mammalian GnT I (designated
gly-12,
gly-13 and
gly-14) have been cloned. All three cDNAs encode proteins with GnT I enzyme activity. We report in this paper the preparation by ultra-violet (UV) light irradiation in the presence of trimethylpsoralen, of mutants lacking either
gly-12,
gly-13 or
gly-14. A double null mutation in the
gly-12 and
gly-14 genes (
gly-14;
gly-12) has also been prepared. These mutations are intragene deletions, removing large portions of the GnT I catalytic domain, and are therefore, all molecular nulls. The
gly-12 and
gly-14 mutants as well as the
gly-14;
gly-12 double mutant all displayed wild-type phenotypes, indicating that neither
gly-12 nor
gly-14 is necessary for worm development under standard laboratory conditions. In contrast, about 60% of the mutants lacking the
gly-13 gene arrested as L1 larvae at 20 degreesC and the remaining 40% homozygous worms grew to adulthood but displayed severe morphological and behavioral defects despite the presence of the other two GnT I genes,
gly-12 and
gly-14. Attempts to rescue the
gly-13 null phenotype with the wild type transgene were not successful. However, lethality co-segregated with the
gly-13 deletion within 0.02 map units (mu) in genetic mapping experiments, suggesting that the
gly-13 mutation is responsible for the phenotype.