Molecular cloning and sequence analysis of an ascidian egg beta-N-acetylhexosaminidase with a potential role in fertilization.
Beta-N-acetylhexosaminidase, which is found almost ubiquitously in sperm of invertebrates and vertebrates, supposedly mediates a carbohydrate-based transient sperm-egg coat binding. In ascidians and mammals, beta-hexosaminidase released at fertilization from eggs has been proposed to modify sperm receptor glycoproteins of the egg envelope, thus setting up a block to polyspermy. Previously, it was shown that in potential sperm receptor glycoproteins of the ascidian Phallusia mammillata, N-acetylglucosamine is the prevailing glycoside residue and that the egg harbors three active molecular forms of beta-hexosaminidase. In the present study, P. mammillata beta-hexosaminidase cDNA was isolated from an ovarian cDNA library and characterized. The deduced amino acid sequence showed a high similarity with other known beta-hexosaminidases; however, P. mammillata beta-hexosaminidase had a unique potential N-glycosylation site. A phylogenetic analysis suggested that P. mammillata beta-hexosaminidase developed independently after having branched off from the common ancestor gene of the chordate enzyme before two isoforms of the mammalian enzyme appeared. In situ hybridization revealed stage-specific expression of beta-hexosaminidase mRNA during oogenesis in the oocyte and in the accessory test and follicle cells. This suggests that the three egg beta-hexosaminidase forms are specific for the oocyte, test cells and follicle cells.
Koyanagi
Ryo
R
Zoological Institute, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.
Honegger
Thomas G
TG
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D017931:DNA Primers; D005260:Female; D005306:Fertilization / Q000235:genetics*; D005602:France; D015723:Gene Library; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D010063:Ovum / Q000737:chemistry*; D010802:Phylogeny; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D014561:Urochordata / Q000235:genetics*; D001619:beta-N-Acetylhexosaminidases / Q000235:genetics*
D017931:DNA Primers; D001619:beta-N-Acetylhexosaminidases