PKC isotypes in post-activated and fertilized mouse eggs: Association with the meiotic spindle

Several isotypes of protein kinase C (PKC) have been reported to be expressed in mammalian eggs, but it is unknown whether these isotypes have a common function in the egg during or within the first few hours of fertilization. Here we show that the isotypes of PKC exhibit distinct patterns of enrichment immediately after mouse egg activation. PKCα and γ accumulate in the egg cortex 25 min post-activation, while only PKCα accumulates at the contractile ring of the forming second polar body about 1.5 h post-activation. PKCζ exhibits some unique features that resulted in it being the focus of more extensive analysis. PKCζ is tightly associated with the meiotic spindle as determined by detergent extraction and is closely associated with α-tubulin as determined by FRET analysis in the metaphase II (MII) egg. In addition, after egg activation, PKCζ remains associated with the spindle as it transits into anaphase II and later telophase II, becoming associated with the midzone microtubules. Antibodies to the active form of PKCζ are enriched on the spindle poles and later in development on the midzone microtubules. Active PKCζ also is enriched in both pronuclei in the 6-h post-fertilization and in the 14-h post-fertilization embryo as well as in the nuclei of the two-cell embryo. Inhibition of PKCζ, but not inhibition of other isotypes of PKC, results in rapid disruption of the meiotic spindle. This study suggests that PKCζ has a role in spindle stability, while other PKC isotypes have different roles in the conversion of the egg to the zygote.