Identification and expression analysis of Drosophila melanogaster genes encoding β-hexosaminidases of the sperm plasma membrane

Sperm surface β-N-acetylhexosaminidases are among the molecules mediating early gamete interactions in invertebrates and vertebrates, including man. The plasma membrane of Drosophila spermatozoa contains two β-N-acetylhexosaminidases, DmHEXA and DmHEXB, which are required for egg fertilization. Here, we demonstrate that three putative Drosophila melanogaster genes predicted to code for β-N-acetylhexosaminidases, Hexo1, Hexo2, and fdl, are all expressed in the male germ line. fdl codes for a homolog of the α-subunit of the mammalian lysosomal β-N-acetylhexosaminidase Hex A. Hexo1 and Hexo2 encode two homologs of the β-subunit of all known β-N-acetylhexosaminidases, which we have named β₁ and β₂, respectively. Immunoblot analysis of sperm proteins indicated that the gene products associate in different heterodimeric combinations forming DmHEXA, with an αβ₂ structure, and DmHEXB, with a β₁ β₂ structure. Immunofluorescence demonstrated that all the gene products localized to the sperm plasma membrane. Although none of the genes was testis-specific, fdl was highly and preferentially expressed in the testis, whereas Hexo1 and Hexo2 showed broader tissue expression. Enzyme assays carried out on testis and on a variety of somatic tissues corroborated the results of gene expression analysis. These findings for the first time show the in vivo expression in insects of genes encoding β-N-acetylhexosaminidases, the only molecules so far identified as involved in sperm/egg recognition in this class, whereas in mammals, the organisms where these enzymes have been best studied, only two types of polypeptide chains forming dimeric functional β-N-acetylhexosaminidases are present in Drosophila three different gene products are available that might generate numerous dimeric isoforms.