A previous screen for C. elegans mutants defective in vulval invagination identified eight sqv genes (squashed vulva). In sqv mutant L4 larvae the vulval extracellular space fails to expand and the vulva appears collapsed. Stronger sqv mutations also show maternal effect lethality as a result of the inability of the progeny of homozygous mutants to initiate cytokinesis at the first cell division. Characterization of seven sqv genes showed they define components of the glycosaminoglycan biosynthesis pathway. Three of these gene products generate or transport nucleotide sugar precursors: SQV-4 is the UDP-glucose dehydrogenase that catalyzes formation of UDP-glucuronic acid; SQV-1 is a decarboxylase that synthesizes UDP-xylose; and SQV-7 is a multi-transporter that imports three nucleotide sugars into the Golgi apparatus. Studies of SQV-6, SQV-3, SQV-2, and SQV-8 identified them as xylosyltransferase, galactosyltransferase-I, galactosyltransferase-II, and glucuronosyltransferase-I, respectively. These enzymes are required for the assembly of the proteoglycan core protein linkage tetrasaccharide GlcA1,3Gal1,3Gal1,4Xyl-O-(Ser), which is common to the two major types of glycosaminoglycans, chondroitin and heparan sulfate. We report that the eighth sqv gene,
sqv-5, is the C. elegans ortholog of the chondroitin synthase involved in polymerization of the chondroitin chain. Biochemical assays showed substantial synthase activity in wild-type worms, approximately one-half of that activity in
sqv-5 heterozygotes, and virtually no activity in homozygous mutants. In contrast, activity levels of the heparan sulfate co-polymerase were comparable in wild-type and
sqv-5 mutant worms. Chemical analyses demonstrated that
sqv-5 mutants have negligible chondroitin levels, while wild-type worms contain 180 fmol/worm. Western blots of partially purified proteoglycans showed that worms treated with
sqv-5 RNAi had a reduction in chondroitin-containing proteoglycans, while levels of heparan sulfate proteoglycans remained constant. Because the phenotype of the
sqv-5 mutant appears to be identical to that of the other sqv mutants, these data suggest that vulval and embryonic defects are a consequence of a lack of chondroitin, not heparan sulfate, in the developing animal. Studies are underway to identify the major proteoglycan core protein involved in these processes.