The role of PAR-3 (partitioning defective gene-3) in the development of asymmetry during mouse meiotic maturation

During mammalian oocyte maturation, the radially symmetric oocyte is transformed into a highly polarized metaphase II (MII)-arrested egg. How this polarity is established is unknown. The evolutionarily conserved partitioning defective (PAR) proteins are pivotal for regulating polarity in numerous cell types. To determine if PAR-3 could be involved in mammalian oocyte maturation, several aspects of PAR-3 expression were examined. Two novel Par-3 transcripts were cloned from an oocyte cDNA library, which encode proteins that are phosphorylated during maturation. PAR-3 becomes asymmetrically localized during meiosis. Following germinal vesicle breakdown, PAR-3 surrounds the condensing chromosomes and associates with the meiotic spindles. Prior to emission of the first and second polar bodies, PAR-3 is located within a central subdomain of the polarized actin cap, overlying the spindle. This cortical PAR-3 localization depends on intact microfilaments. These results suggest a role for PAR-3 in establishing asymmetry in the egg and in defining the site of polar body emission. To test this hypothesis, several approaches were used to attempt to disrupt PAR-3 function, including antibody microinjection, and overexpression and mislocalization of PAR-3 isoforms. These approaches did not affect the ability of oocytes to resume meiosis, form an actin cap, or extrude the first polar body. To determine the effect of PAR-3 loss-of-function, a transgenic RNA interference approach was used. The ZP3 promoter was used to drive expression of a hairpin dsRNA targeting Par-3 mRNA in growing oocytes. The transgene resulted in a Par-3 mRNA knockdown of up to 90% which corresponded to a 50% protein reduction. Founder female fertility appeared normal, but transgene mosaicism in the germline and a modest protein decrease may have masked a subfertility phenotype. Although these functional studies suggest that PAR-3 is not the critical asymmetry regulator in mouse oocytes, whether the approaches used adequately disrupted function is unknown. Future studies to knockout Par-3, determine PAR-3-interacting proteins, and examine other asymmetry regulators are warranted to understand how polarity is established during meiosis.