Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki-, Japan.
Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki-, Japan.
Biol Reprod. 2019 Jun 1;100(6):1482-1491. doi: 10.1093/biolre/ioz015.
Despite knowledge that glucose metabolism is essential for the regulation of signaling cascades in the sperm that are pre-assembled into specific areas and function at multistage for fertilization, the physiological roles of glucose in avian sperm are poorly understood. Accumulated results of studies conducted in our laboratory and others indicate that sperm possess membrane microdomains, or membrane rafts, which play important roles in several processes, including the induction of acrosome reaction (AR). When characterizing proteomes associated with chicken sperm rafts, we observed marked enrichment of glucose transporter 3 (GLUT3). Here we show that glucose uptake is mediated by membrane rafts and stimulates AR induction by activating AMP-activated protein kinase (AMPK). Using a specific antibody, we observed that GLUT3 is localized to the entire flagellum and acrosome region and highly associated with membrane rafts. The addition of glucose stimulated AR in a dose-dependent manner without affecting sperm motility. AR and glucose uptake assays were performed using both inhibitors and activators, and demonstrated that glucose-dependent AR results from the activity of a glucose transporter located in membrane rafts and associated with AMPK. To better understand the mechanism of AMPK activation by glucose, we evaluated localization and phosphorylation status of AMPKα, showing that glucose uptake stimulates AMPKα phosphorylation, leading to its complete activation. Together, these results lead us to propose a novel mechanism by which glucose uptake stimulates the AMPK signaling pathway via membrane rafts, resulting in maximal acrosomal responsiveness in avian sperm as migrating upward to a fertilization site.
尽管人们知道葡萄糖代谢对于精子中信号级联的调节至关重要，这些信号级联预先组装到特定区域，并在多阶段受精过程中发挥作用，但葡萄糖在禽类精子中的生理作用仍知之甚少。我们实验室和其他实验室的研究结果表明，精子具有膜微区或膜筏，它们在包括顶体反应（AR）诱导在内的几个过程中发挥重要作用。在对与鸡精子筏相关的蛋白质组进行特征描述时，我们观察到葡萄糖转运蛋白 3（GLUT3）明显富集。在这里，我们表明葡萄糖摄取是通过膜筏介导的，并通过激活 AMP 激活的蛋白激酶（AMPK）来刺激 AR 诱导。使用特异性抗体，我们观察到 GLUT3 定位于整个鞭毛和顶体区域，并与膜筏高度相关。添加葡萄糖以剂量依赖的方式刺激 AR，但不影响精子活力。使用抑制剂和激活剂进行了 AR 和葡萄糖摄取测定，表明葡萄糖依赖性 AR 是由位于膜筏中并与 AMPK 相关的葡萄糖转运体的活性引起的。为了更好地理解葡萄糖对 AMPK 激活的机制，我们评估了 AMPKα的定位和磷酸化状态，结果表明葡萄糖摄取刺激 AMPKα磷酸化，导致其完全激活。总之，这些结果使我们提出了一种新的机制，即葡萄糖通过膜筏摄取刺激 AMPK 信号通路，从而导致禽类精子顶体反应达到最大，因为它们向上迁移到受精部位。
