The pattern of localization of the putative oocyte activation factor, phospholipase Czeta, 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, PLCzeta. We investigated PLCzeta localization in human spermatozoa.


METHODS
A polyclonal antibody was generated against human PLCzeta 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-PLCzeta antibody by immunoblotting, immunofluorescence studies showed that the predominant localization of PLCzeta in uncapacitated sperm was in the equatorial region, a pattern maintained following capacitation and ionophore treatment. The analysis of pooled samples showed approximately 88% of uncapacitated sperm expressed PLCzeta in the equatorial region, whereas approximately 35% and approximately 21% of sperm expressed additional populations of PLCzeta in the acrosomal or post-acrosomal region, respectively. One population of PLCzeta was observed in the post-acrosomal region of approximately 12% of sperm. The proportion of cells with post-acrosomal PLCzeta 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 PLCzeta in the acrosomal region.


CONCLUSIONS
PLCzeta was variably detectable in three localities within the sperm head: the equatorial segment and acrosomal/post-acrosomal region. Variability in PLCzeta 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 PLCzeta and certain types of male infertility.
Grasa
P
P
Department of Pharmacology, University of Oxford, Oxford, UK.
Coward
K
K
Young
C
C
Parrington
J
J
D000177:Acrosome / Q000378:metabolism; D005260:Female; D006801:Humans; D007248:Infertility, Male / Q000188:drug therapy / Q000209:etiology; D007476:Ionophores / Q000494:pharmacology*; D008297:Male; D008856:Microscopy, Fluorescence / Q000379:methods; D009865:Oocytes / Q000187:drug effects* / Q000378:metabolism*; D054801:Phosphoinositide Phospholipase C / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry; D012680:Sensitivity and Specificity; D013075:Sperm Capacitation; D013081:Sperm Motility; D013094:Spermatozoa / Q000187:drug effects* / Q000378:metabolism*
D007476:Ionophores; D011994:Recombinant Proteins; C499235:PLCZ1 protein, human; D054801:Phosphoinositide Phospholipase C