Meiotic induction by heat stress in mouse oocytes: Involvement of AMP-activated protein kinase and MAPK family members
Meiotic induction by heat stress in mouse oocytes: Involvement of AMP-activated protein kinase and MAPK family members
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DOI：
10.1095/biolreprod.106.057422
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发表时间：
2007-03-01
影响因子：
3.6
通讯作者：
Downs, Stephen M.
中科院分区：
文献类型：
--
作者：
LaRosa, Cean;Downs, Stephen M.
在这项研究中，我们研究了热脉冲对卵母细胞成熟的影响，并评估了热应激诱导减数分裂成熟过程中应激激活酶的可能作用。将来自未成熟的eCG致敏小鼠的裸露卵母细胞在含二丁酰cAMP的培养基中在40 ℃至43 ℃的温度下脉冲30分钟，随后在37 ℃下培养17-18小时的总孵育时间。暴露于42摄氏度的钟细胞显示出最大的成熟刺激，在43摄氏度没有影响。通过极体（PB）形成观察到，热脉冲不会影响向中期II的进展。AMP活化蛋白激酶（PRKA）抑制剂化合物C和Ara-A各自阻断热的减数分裂刺激作用。蛋白质印迹显示，乙酰辅酶A羧化酶，一个重要的底物PRKA，被磷酸化热处理的germinal囊泡阶段的卵母细胞，表明激活的PRKA成熟前。丝裂原活化蛋白2激酶（MAP 2K 1）抑制剂PD 98059也防止热诱导成熟，但这种效果是无关的MAPK 1/3激活，这是没有观察到，直到后germinal囊泡破裂（GVB）。在任何实验条件下，在卵母细胞中均未检测到磷酸化MAPK 14，并且仅高浓度的MAPK 14抑制剂SR 203580阻断热刺激成熟，表明MAPK 14不参与减数分裂诱导。MAPK 8/9被热激活，MAPK 8/9抑制剂SP 600125，但不是JUN N-末端激酶1，阻断热诱导的成熟。热处理瞬时抑制自发成熟卵母细胞的GVB和PB形成的机制，这显然是不同的减数分裂诱导作用。总的来说，这些数据表明，急性热脉冲刺激GVB减数分裂停滞的卵母细胞，并表明这种效果是通过激活PRKA介导的。
In this study, we examined the effect of heat pulsing on oocyte maturation and assessed the possible role of stress-activated enzymes during heat stress-induced meiotic maturation. Denuded oocytes from immature eCG-primed mice were pulsed for 30 min at increasing temperatures from 40 degrees C to 43 degrees C in dibutyryl cAMP-containing medium and were subsequently cultured at 37 degrees C for a total incubation time of 17-18 h. Clocytes exposed to 42 degrees C showed the greatest stimulation of maturation, with no effect at 43 degrees C. A heat pulse did not compromise progression to metaphase II as observed by polar body (PB) formation. The AMP-activated protein kinase (PRKA) inhibitors compound C and Ara-A each blocked the meiosis-stimulating effects of heat. Western blots showed that acetyl-CoA carboxylase, an important substrate of PRKA, was phosphorylated in heat-treated germinal vesicle-stage oocytes, indicating activation of PRKA before maturation. The mitogen-activated protein 2 kinase (MAP2K1) inhibitor PD98059 also prevented heat-induced maturation, but this effect was unrelated to MAPK1/3 activation, which was not observed until after germinal vesicle breakdown (GVB). Phosphorylated MAPK14 was not detected in the oocyte under any experimental condition, and only high concentrations of the MAPK14 inhibitor SR203580 blocked heat-stimulated maturation, suggesting that MAPK14 is not involved in meiotic induction. MAPK8/9 was activated by heat, and the MAPK8/9 inhibitor SP600125, but not JUN N-terminal kinase 1, blocked heat-induced maturation. Heat treatment transiently suppressed GVB and PB formation in spontaneously maturing oocytes by a mechanism that is apparently different from its meiosis-inducing action. Collectively, these data show that an acute heat pulse stimulates GVB in meiotically arrested oocytes and suggest that this effect is mediated through the activation of PRKA.