Brandelli A, Miranda P V, Tezón J G
Departamento de Bioquimica, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil.
J Androl. 1996 Sep-Oct;17(5):522-9.
Mammalian spermatozoa must undergo an exocytotic event during fertilization, the acrosome reaction (AR). In most species studied this process is induced by specific glycoproteins of the oocyte extracellular matrix, the zona pellucida (ZP), and it involves guanine nucleotide-binding regulatory proteins (G-proteins), resulting in an uptake of extracellular calcium by the sperm. In the bull, this event has been reported to be mediated by voltage-dependent calcium channels (VDCC). Previous observations showed that neoglycoproteins (NGPs) with N-acetylglucosamine or mannose (GlcNAc-BSA or Man-BSA) residues induce the AR in capacitated human spermatozoa. We report here that the pretreatment of spermatozoa with 125 ng/ml pertussis toxin (PTx) inhibited GlcNAc-BSA- or Man-BSA-induced AR, whereas 1 microgram/ml cholera toxin had no effect. These data indicate that the transduction mechanism for GlcNAc-BSA- and Man-BSA-induced AR involves G-proteins of the inhibitory type (GI). An increase in the AR rate was observed when capacitated spermatozoa were incubated with increasing concentrations of potassium ions (K+) in Biggers-Whitten-Whittingham (BWW) modified medium (2.6 +/- 0.3-fold at 80 mM K+). This induction was observed only when the pH was raised to 8.5, and it was inhibited by verapamil, nitrendipine, omega-conotoxin, nickel ions (Ni2+), lanthanum ions (La3+), or cadmium ions (Cd2+) in a concentration-dependent manner, indicating the participation of VDCC activated by membrane depolarization. The GlcNAc-BSA- or Man-BSA-induced AR was completely inhibited by preincubation of spermatozoa with VDCC blockers and calcium antagonists, indicating a link between the binding of sugar residues of the NGPs and channel activation. The AR induced by membrane depolarization with high K+ medium was not inhibited by PTx, suggesting that Ca2+ entry is downstream to GI-protein activation. These data show that the induction of the AR in human spermatozoa by GlcNAc- or Man-NGPs involves VDCC and GI-like regulatory proteins similar to the induction described for ZP in other mammalian species.
哺乳动物的精子在受精过程中必须经历一种胞吐事件，即顶体反应（AR）。在大多数已研究的物种中，这个过程是由卵母细胞细胞外基质——透明带（ZP）的特定糖蛋白诱导的，并且涉及鸟嘌呤核苷酸结合调节蛋白（G蛋白），导致精子摄取细胞外钙。在公牛中，据报道这一事件是由电压依赖性钙通道（VDCC）介导的。先前的观察表明，带有N - 乙酰葡糖胺或甘露糖（GlcNAc - BSA或Man - BSA）残基的新糖蛋白（NGP）可诱导获能的人类精子发生顶体反应。我们在此报告，用125 ng/ml百日咳毒素（PTx）预处理精子可抑制GlcNAc - BSA或Man - BSA诱导的顶体反应，而1 μg/ml霍乱毒素则无此作用。这些数据表明，GlcNAc - BSA和Man - BSA诱导顶体反应的转导机制涉及抑制型G蛋白（GI）。当获能的精子在改良的Biggers - Whitten - Whittingham（BWW）培养基中与浓度不断增加的钾离子（K +）一起孵育时，观察到顶体反应率增加（在80 mM K +时为2.6±0.3倍）。仅当pH值升至8.5时才观察到这种诱导作用，并且它被维拉帕米、尼群地平、ω - 芋螺毒素、镍离子（Ni2 +）、镧离子（La3 +）或镉离子（Cd2 +）以浓度依赖性方式抑制，表明膜去极化激活的VDCC参与其中。精子与VDCC阻滞剂和钙拮抗剂预孵育可完全抑制GlcNAc - BSA或Man - BSA诱导顶体反应，这表明NGP糖残基的结合与通道激活之间存在联系。高钾培养基膜去极化诱导的顶体反应不受PTx抑制，这表明Ca2 +内流是在GI蛋白激活的下游。这些数据表明，GlcNAc或Man - NGP诱导人类精子发生顶体反应涉及VDCC和类似于其他哺乳动物物种中ZP诱导所描述的GI样调节蛋白。
