Involvement of ER-calreticulin-Ca²⁺ signaling in the regulation of porcine oocyte meiotic maturation and maternal gene expression

Calcium is one of themost ubiquitous signaling molecules, and controls awide variety of cellular processes. It is mainly stored in the endoplasmic reticulum (ER), bound to lumenal proteins. Calreticulin is the major Ca ²⁺-binding chaperone in oocytes, and is integral to numerous cellular functions. To better understand the role of the ER-calreticulin-Ca²⁺ pathway in oocyte maturation and early embryogenesis, we characterized the porcine calreticulin gene and investigated its expression profile during oocyte maturation and early embryonic development. Calreticulin was widely expressed in pig tissues and its transcripts were downregulated during maturation, especially at 44 hr, and were undetectable at the blastocyst stage. We also investigated the effect of increased cytosolic Ca²⁺ induced by the Ca²⁺-ATPase inhibitor, cyclopiazonic acid (CPA), on pig oocyte maturation and maternal gene expression. CPA at 10 mM did not inhibit germinal vesicle breakdown, but did result in the arrest of 38.6% oocytes at or before the MI stage. In addition, expression of the maternal genes C-mos, BMP15, GDF9, and Cyclin B1 was significantly increased in CPA-treated MII oocytes compared with control groups. These data were supported by the results of poly(A)-test PCR, which revealed that the cyclin B1 short isoform (CB-S), GDF9, and C-mos underwent more intensive polyadenylation modification in CPA-treated oocytes than control oocytes, suggesting that polyadenylation may influence Ca ²⁺-modulated changes in gene expression. Furthermore, CPA treatment decreased the percentage of four-cell parthenotes that developed into blastocysts, suggesting the need for functional SR/ER Ca²⁺-ATPase pumps or Ca²⁺ signals during early embryo development after zygotic genome activation. Together, these data indicate that ER-calreticulin-associated Ca²⁺ homeostasis plays a role in oocyte and embryo development, and that alterations in maternal gene expression may contribute to the underlying molecular mechanism, at least partially, via polyadenylation.