Kobrinsky E M, Kirchberger M A
Department of Physiology and Biophysics, The Mount Sinai School of Medicine of New York University, New York, NY 10029, USA.
Oncogene. 2001 Feb 22;20(8):933-41. doi: 10.1038/sj.onc.1204153.
Thapsigargin (Tg), a selective inhibitor of sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase (SERCA), causes depletion of intracellular Ca(2+) stores, hence activation of capacitative Ca(2+) entry (CCE). Incubation of Xenopus laevis oocytes with Tg resulted in an increased rate of progesterone-induced meiotic maturation. Non-mitochondrial (45)Ca(2+) uptake by SERCA-containing microsomes prepared from control wild-type oocytes microinjected with sterile water was inhibited essentially 100% by Tg. However, overexpression of Bcl-2, an oncogene known to protect against Tg-induced apoptosis in certain cell types, resulted in only 40% inhibition of microsomal (45)Ca(2+) uptake by Tg while non-inhibited (45)Ca(2+) uptake remained unchanged. Moreover Bcl-2 overexpression also protected against inhibition of CCE. I(Cl(Ca)) was similar in Bcl-2-overexpressing and control oocytes when intracellular Ca(2+) store depletion was induced by microinjection of inositol 1,4,5-trisphosphate (InsP(3)) and other means and when CCE was induced by means independent of SERCA inhibition. Our data indicate that Bcl-2 affects neither the InsP(3) receptor nor Ca(2+) entry itself. At the end of a 24-h period after progesterone addition to the medium, only 25% of Bcl-2-overexpressing oocytes had matured compared to 85% of control oocytes. Our data suggest that SERCA participates in Xenopus oocyte maturation by controlling cytosolic Ca(2+) and/or intracellular Ca(2+) stores, hence CCE. An observed progesterone-dependent protein kinase-catalysed phosphorylation of SERCA is further indication of its role in oocyte maturation.
毒胡萝卜素（Tg）是肌浆网/内质网Ca²⁺-ATP酶（SERCA）的选择性抑制剂，可导致细胞内Ca²⁺储存耗尽，从而激活容量性Ca²⁺内流（CCE）。用Tg孵育非洲爪蟾卵母细胞会导致孕酮诱导的减数分裂成熟速率增加。用无菌水显微注射对照野生型卵母细胞制备的含SERCA的微粒体对非线粒体（⁴⁵）Ca²⁺摄取基本上被Tg抑制了100%。然而，Bcl-2（一种已知在某些细胞类型中可防止Tg诱导的细胞凋亡的癌基因）的过表达导致Tg对微粒体（⁴⁵）Ca²⁺摄取的抑制仅为40%，而未受抑制的（⁴⁵）Ca²⁺摄取保持不变。此外，Bcl-2过表达还可防止CCE受到抑制。当通过显微注射肌醇1,4,5-三磷酸（InsP₃）和其他方法诱导细胞内Ca²⁺储存耗尽时，以及当通过独立于SERCA抑制的方法诱导CCE时，Bcl-2过表达的卵母细胞和对照卵母细胞中的I(Cl(Ca))相似。我们的数据表明，Bcl-2既不影响InsP₃受体，也不影响Ca²⁺内流本身。在向培养基中添加孕酮24小时后，只有25%的Bcl-2过表达卵母细胞成熟，而对照卵母细胞为85%。我们的数据表明，SERCA通过控制胞质Ca²⁺和/或细胞内Ca²⁺储存，从而参与非洲爪蟾卵母细胞的成熟，进而参与CCE。观察到的孕酮依赖性蛋白激酶催化的SERCA磷酸化进一步表明了其在卵母细胞成熟中的作用。
