TARGETING OF A GERM CELL-SPECIFIC TYPE-1 HEXOKINASE LACKING A PORIN-BINDING DOMAIN TO THE MITOCHONDRIA AS WELL AS TO THE HEAD AND FIBROUS SHEATH OF MURINE SPERMATOZOA

Multiple isoforms of type 1 hexokinase (HK1) are transcribed during sp ermatogenesis in the mouse, including at least three that are presumab ly germ cell specific: HK1-sa, HK1-sb, and HK1-sc. Each of these predi cted proteins contains a common, germ cell-specific sequence that repl aces the porin-binding domain found in somatic HK1. Although HK1 prote in is present in mature sperm and is tyrosine phosphorylated, it is no t known whether the various potential isoforms are differentially tran slated and localized within the developing germ cells and mature sperm . Using antipeptide antisera against unique regions of HK1-sa and HK1- sb, it was demonstrated that these isoforms were not found in pachyten e spermatocytes, round spermatids, condensing spermatids, or sperm, su ggesting that HK1-sa and HK1-sb are not translated during spermatogene sis. Immunoreactivity was detected in protein from round spermatids, c ondensing spermatids, and mature sperm using an antipeptide antiserum against the common, germ cell-specific region, suggesting that HK1-sc was the only germ cell-specific isoform present in these cells. Two-di mensional SDS-PAGE suggested that all of the sperm HK1-sc was tyrosine phosphorylated, and that the somatic HK1 isoform was not present. Imm unoelectron microscopy revealed that HK1-sc was associated with the mi tochondria and with the fibrous sheath of the flagellum and was found in discrete clusters in the region of the membranes of the sperm head. The unusual distribution of HK1-sc in sperm suggests novel functions, such as extramitochondrial energy production, and also demonstrates t hat a hexokinase without a classical porin-binding domain can localize to mitochondria.