Egg shell collagen formation in Caenorhabditis elegans involves a novel prolyl 4-hydroxylase expressed in spermatheca and embryos and possessing many unique properties.
The collagen prolyl 4-hydroxylases (EC ) play a critical role in the synthesis of all collagens. The enzymes from all vertebrate species studied are alpha(2)beta(2) tetramers, in which the beta subunit is identical to protein disulfide isomerase (PDI). Two isoforms of the catalytic alpha subunit, PHY-1 and PHY-2, have previously been characterized from Caenorhabditis elegans. We report here on the cloning and characterization of a third C. elegans alpha subunit isoform, PHY-3. It is much shorter than the previously characterized vertebrate and C. elegans alpha subunits and shows 23-30% amino acid sequence identity to PHY-1 and PHY-2 within the catalytic C-terminal region. Recombinant PHY-3 coexpressed in insect cells with a C. elegans PDI isoform that does not associate with PHY-1 was found to be an active prolyl 4-hydroxylase. The phy-3 gene consists of five exons, and its expression pattern differs distinctly from the hypodermally expressed phy-1 and phy-2 in that it is expressed in embryos, late larval stages, and adult nematodes, expression in the latter being restricted to the spermatheca. Nematodes homozygous for a phy-3 deletion are phenotypically of the wild type and fertile, but the 4-hydroxyproline content of phy-3(-/-) early embryos was reduced by about 90%. PHY-3 is thus likely to be involved in the synthesis of collagens in early embryos, probably of those in the egg shell.
Riihimaa
Paivi
P
Collagen Research Unit, Biocenter Oulu and Department of Medical Biochemistry, University of Oulu, FIN-90014 Oulu, Finland.
Nissi
Ritva
R
Page
Antony P
AP
Winter
Alan D
AD
Keskiaho
Katriina
K
Kivirikko
Kari I
KI
Myllyharju
Johanna
J
D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology* / Q000254:growth & development; D020134:Catalytic Domain; D003001:Cloning, Molecular; D003094:Collagen / Q000096:biosynthesis*; D004528:Egg Shell; D006801:Humans; D006909:Hydroxyproline; D007527:Isoenzymes / Q000378:metabolism; D008969:Molecular Sequence Data; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism*; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D013997:Time Factors
D007527:Isoenzymes; D003094:Collagen; D011393:Procollagen-Proline Dioxygenase; D019704:Protein Disulfide-Isomerases; D006909:Hydroxyproline