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当前位置: 首页 > 文章 > 钙离子对猪精子活力及蛋白磷酸化调节作用研究 畜牧与兽医 2015,47 (9) 20-26
Position: Home > Articles > Regulatory roles of calcium on sperm motility and protein phosphorylation in boars Animal Husbandry & Veterinary Medicine 2015,47 (9) 20-26

钙离子对猪精子活力及蛋白磷酸化调节作用研究

作  者：
赵娜;甄林青;王勃;陆林根;李新红
单  位：
上海交通大学农业与生物学院;上海市嘉定区梅山猪育种中心
关键词：
猪精子;获能;蛋白磷酸化;钙离子;精子活力
摘  要：
钙离子(Ca2+)参与哺乳动物精子获能、超激活运动和顶体反应等多个生理过程,但其作用机理尚未明确。本研究旨在探讨Ca2+在猪精子获能过程中对精子活力和蛋白磷酸化的影响及其作用机制。采用计算机辅助精子活力分析仪(CASA)及蛋白免疫印迹方法测定不同Ca2+浓度处理后精子活力参数及蛋白磷酸化水平变化,同时,利用细胞免疫荧光技术对不同处理组精子样品磷酸化蛋白进行细胞亚组分定位。CASA结果显示,随着Ca2+浓度增加,猪精子活力参数呈现先增加(<1.0 mmol/L)后降低(>3.0 mmol/L)的趋势;免疫印迹结果发现,低浓度钙离子(0.5 mmol/L)促进精子蛋白磷酸化,而高浓度钙离子(4.0 mmol/L)显著抑制蛋白磷酸化;免疫荧光揭示,4.0 mmol/L钙离子处理组鞭毛处磷酸化蛋白明显少于未获能组和获能组;c AMP-PKA信号通路调节因子c AMP和IBMX以及钙调蛋白抑制剂W7和CZ均能有效缓解高浓度钙离子对猪精子蛋白酪氨酸磷酸化的抑制作用。结果提示,钙离子对猪精子活力及蛋白磷酸化起双重调节作用:即低浓度钙离子(0.5 mmol/L)有助于提高猪精子活力及蛋白磷酸化,高浓度钙离子(4.0 mmol/L)抑制精子活力和蛋白磷酸化;而高浓度钙离子(4.0 mmol/L)可能通过c AMPPKA信号通路和Ca2+/Ca M通路抑制鞭毛处与精子活力有关的蛋白(动力蛋白或轴丝蛋白)磷酸化,进而抑制精子活力。
译  名：
Regulatory roles of calcium on sperm motility and protein phosphorylation in boars
作  者：
ZHAO Na;ZHEN Linqing;WANG Bo;LU Lingen;LI Xinhong;Shanghai Key Laboratory of Veterinary Biotechnology,College of Agriculture and Biology,Shanghai Jiaotong University;Meishan Boar Breeding Center of Jiading;
关键词：
boar sperm;;capacitation;;protein phosphorylation;;Ca2 +;;sperm motility
摘  要：
In spite of the importance of Ca2 + in regulating sperm capacitation,hyperactivation and acrosome reaction,little is known about the mechanism that regulate these physiological and biochemical processes. The objective of the present work was to investigate the mechanism of Ca2 +regulation on sperm motility and protein phosphorylation in boars. Sperm motility parameters were determined using computerassisted sperm analysis( CASA) and the phosphorylated status of PKA substrates and tyrosine residueswere analyzed by western blotting.Meanwhile,localization of phosphorylated protein targets in the boar sperm was observed using immunofluorescence technique. Our results indicated that sperm motility was significantly decreased and protein phosphorylation was inhibited under high extracellular Ca2 +concentration( 4. 0 mmol/L). In addition,phosphorylated protein substrates with 4. 0 mmol/L Ca2 +treatment were significantly less than N- Cap and Cap treatments. Interestingly,the negative effect of Ca2 +( 4. 0 mmol/L) was overcome by addition of c AMP,IBMX,calmodulin inhibitors,W7 and CZ. Taken together,our results indicate that changes in extracellular Ca2 +play a biphasic role in the regulation of sperm motility and protein phosphorylation. A certain concentration of calcium( 0. 5 mmol / L) plays a promoting effect on boar sperm capacitation and sperm motility,but the higher concentration of calcium( 4. 0 mmol / L) has a inhibitory effect. Negative effects of Ca2 +( 4. 0 mmol/L) on sperm motility maybe via the c AMP / PKA and Ca2 +/ Ca M pathways to suppress protein phosphorylation of dynein and axokinin in the flagella.

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