Tyrosine phosphorylation of the A kinase anchoring protein 3 (AKAP3) and soluble adenylate cyclase are involved in the increase of human sperm motility by bicarbonate
Tyrosine phosphorylation of the A kinase anchoring protein 3 (AKAP3) and soluble adenylate cyclase are involved in the increase of human sperm motility by bicarbonate
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DOI：
10.1095/biolreprod.104.032490
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发表时间：
2005-01-01
影响因子：
3.6
通讯作者：
Baldi, E
中科院分区：
文献类型：
--
作者：
Luconi, M;Porazzi, I;Baldi, E
哺乳动物睾丸精子是不动的，因此，为了到达卵母细胞，它们需要在精子通过附睾和女性生殖道过程中的不同因素的控制下获得游泳能力。尽管已知碳酸氢盐通过刺激哺乳动物精子的可溶性腺苷酸环化酶（sAC）活性而在生理上增加活力，但尚未对人类精子进行广泛的研究来阐明所涉及的其他分子机制。有鉴于此，我们研究了体外向人类精子添加碳酸氢盐对参与运动调节的主要细胞内信号传导途径（即细胞内 cAMP 产生和蛋白质酪氨酸磷酸化）的影响。将碳酸氢盐的作用与磷脂酰肌醇 3 激酶抑制剂 LY294002 的作用进行了比较，LY294002 先前被证明是精子活力的药理学刺激剂。向精子中添加碳酸氢盐会导致精子活力以及一些过度激活参数显着增加。碳酸氢盐和 LY294002 的这种刺激作用是通过 cAMP 产生的增加和 A 激酶锚定蛋白 AKAP3 的酪氨酸磷酸化介导的。碳酸氢盐作用的特异性通过 4,4'-二异硫氰芪-2,2'-二磺酸的抑制得到证实。我们注意到，在人类精子中，碳酸氢盐主要通过激活 sAC 来刺激 AKAP3 酪氨酸磷酸化和精子活力，因为这两种作用都会被 sAC 抑制剂 20H-雌二醇削弱。总之，我们的数据提供了第一个证据，表明碳酸氢盐通过激活 sAC 和 AKAP3 的酪氨酸磷酸化来刺激人类精子活力和过度激活，最终导致 PKA 向 AKAP3 的募集增加。
Mammalian testicular spermatozoa are immotile, thus, to reach the oocyte, they need to acquire swimming ability under the control of different factors acting during the sperm transit through the epididymis and the female genital tract. Although bicarbonate is known to physiologically increase motility by stimulating soluble adenylate cyclase (sAC) activity of mammalian spermatozoa, no extensive studies in human sperm have been performed yet to elucidate the additional molecular mechanisms involved. In this light, we investigated the effect of in vitro addition of bicarbonate to human spermatozoa on the main intracellular signaling pathways involved in regulation of motility, namely, intracellular cAMP production and protein tyrosine phosphorylation. Bicarbonate effects were compared with those of the phosphatidyl-inositol-3 kinase inhibitor, LY294002, previously demonstrated to be a pharmacological stimulus for sperm motility. Bicarbonate addition to spermatozoa results in a significant increase in sperm motility as well as in several hyperactivation parameters. This stimulatory effect of bicarbonate and LY294002 is mediated by an increase in cAMP production and tyrosine phosphorylation of the A kinase anchoring protein, AKAP3. The specificity of bicarbonate effects was confirmed by inhibition with 4,4'-di-isothiocyanostilbene-2,2'-disulfonic acid. We remark that, in human spermatozoa, bicarbonate acts primarily through activation of sAC to stimulate tyrosine phosphorylation of AKAP3 and sperm motility because both effects are blunted by the sAC inhibitor 20H-estradiol. In conclusion, our data provide the first evidence that bicarbonate stimulates human sperm motility and hyperactivation through activation of sAC and tyrosine phosphorylation of AKAP3, finally leading to an increased recruitment of PKA to AKAP3.