The pattern of localization of the putative oocyte activation factor, phospholipase Cζ, in uncapacitated, capacitated, and ionophore-treated human spermatozoa

BACKGROUND Recent studies suggest that in mammals, oocyte activation at fertilization is triggered by a sperm-specific phospholipase C, PLCζ. We investigated PLCζ localization in human spermatozoa. METHODS A polyclonal antibody was generated against human PLCζ and used in immunoblotting and immunofluorescence studies of ejaculated human sperm in uncapacitated and capacitated states. An ionophore was also used to induce the acrosome reaction in vitro. RESULTS After verifying specificity of the anti-PLCζ antibody by immunoblotting, immunofluorescence studies showed that the predominant localization of PLCζ in uncapacitated sperm was in the equatorial region, a pattern maintained following capacitation and ionophore treatment. The analysis of pooled samples showed ∼88% of uncapacitated sperm expressed PLCζ in the equatorial region, whereas ∼35% and ∼21% of sperm expressed additional populations of PLCζ in the acrosomal or post-acrosomal region, respectively. One population of PLCζ was observed in the post-acrosomal region of ∼12% of sperm. The proportion of cells with post-acrosomal PLCζ increased following capacitation and ionophore treatment (P < 0.05). The same tendency was found in individual samples. There was a strong correlation (r = 0.716, P < 0.0001) between presence of an intact acrosome and proportion of sperm immunoreactive to PLCζ in the acrosomal region. CONCLUSIONS PLCζ was variably detectable in three localities within the sperm head: the equatorial segment and acrosomal/post-acrosomal region. Variability in PLCζ localization in sperm from fertile males may reflect differences in oocyte activation capabilities between individuals or within an ejaculate. This approach may help in investigating the possible links between PLCζ and certain types of male infertility.