Calicin in human sperm fertilizing zona-free hamster eggs in vitro

Abstract

Calicin, a basic cytoskeletal protein has been proposed to be involved in the formation and maintenance of the highly regular organization of the postacrosomal perinuclear theca, the calyx of mammalian spermatozoa. The fate of human sperm calicin in the zona-free hamster egg fertilized in vitro has been analysed at the light and electron microscope level using polyclonal mouse anti-calicin antibodies. Calicin was localized in the postacrosomal dense lamina of the capacitated acrosome-reacted spermatozoa attached on the surface of the egg as well as in the spermatozoa at an early stage of ooplasmic incorporation. As determined by the aid of the DNA-specific fluorescent dye 4,6-diamidino-2-phenylindole (DAPI), the pattern of staining with anti-calicin changed from a funnel-shape into a ring soon after the onset of the nuclear decondensation. Later, no anti-calicin labelling could be detected around the more decondensed sperm nuclei. Because the residual, ring-like accumulation of calicin was associated with the least decondensed chromatin, it appears that the degradation of calicin-containing perinuclear theca is intimately involved with the posteriorly advancing nuclear disintegration. The ring formation also suggests that the calyx of human spermatozoa is not structurally homogeneous at least in terms of sensitivity to ooplasmic degradation. Calicin appears to be unaffected by lytic enzymes of the acrosome. The present study further shows that DAPI can be effectively used to analyse sperm nuclear decondensation in vitro.