Reid Andrew T, Anderson Amanda L, Roman Shaun D, McLaughlin Eileen A, McCluskey Adam, Robinson Phillip J, Aitken R John, Nixon Brett
*School of Environmental and Life Sciences, Discipline of Biological Sciences, and School of Environmental and Life Sciences, Discipline of Chemistry, The University of Newcastle, Callaghan, New South Wales, Australia; and Children's Medical Research Institute, The University of Sydney, Westmead, New South Wales, Australia.
*School of Environmental and Life Sciences, Discipline of Biological Sciences, and School of Environmental and Life Sciences, Discipline of Chemistry, The University of Newcastle, Callaghan, New South Wales, Australia; and Children's Medical Research Institute, The University of Sydney, Westmead, New South Wales, Australia
FASEB J. 2015 Jul;29(7):2872-82. doi: 10.1096/fj.14-265553. Epub 2015 Mar 25.
The dynamin family of GTPases has been implicated as novel regulators of the acrosome reaction, a unique exocytotic event that is essential for fertilization. Dynamin activity during the acrosome reaction is accompanied by phosphorylation of key serine residues. We now tested the hypothesis that glycogen synthase kinase 3 (GSK3) is the protein kinase responsible for dynamin phosphorylation at these phosphosites in mouse spermatozoa. Pharmacologic inhibition of GSK3 in mature mouse spermatozoa (CHIR99021: IC50 = 6.7 nM) led to a significant reduction in dynamin phosphorylation (10.3% vs. 27.3%; P < 0.001), acrosomal exocytosis (9.7% vs. 25.7%; P < 0.01), and in vitro fertilization (53% vs. 100%; P < 0.01). GSK3 was shown to be present in developing germ cells where it colocalized with dynamin in the peri-acrosomal domain. However, additional GSK3 was acquired by maturing mouse spermatozoa within the male reproductive tract, via a novel mechanism involving direct interaction of sperm heads with extracellular structures known as epididymal dense bodies. These data reveal a novel mode for the cellular acquisition of a protein kinase and identify a key role for GSK3 in the regulation of sperm maturation and acrosomal exocytosis.
GTP酶的发动蛋白家族被认为是顶体反应的新型调节因子，顶体反应是一种独特的胞吐事件，对受精至关重要。顶体反应过程中的发动蛋白活性伴随着关键丝氨酸残基的磷酸化。我们现在测试了糖原合酶激酶3（GSK3）是负责小鼠精子中这些磷酸位点上发动蛋白磷酸化的蛋白激酶这一假设。对成熟小鼠精子中的GSK3进行药理抑制（CHIR99021：IC50 = 6.7 nM）导致发动蛋白磷酸化显著降低（10.3% 对 27.3%；P < 0.001）、顶体胞吐作用降低（9.7% 对 25.7%；P < 0.01）以及体外受精率降低（53% 对 100%；P < 0.01）。结果表明，GSK3存在于发育中的生殖细胞中，在顶体周围区域与发动蛋白共定位。然而，成熟的小鼠精子在雄性生殖道内通过一种新机制获得了额外的GSK3，该机制涉及精子头部与称为附睾致密小体的细胞外结构直接相互作用。这些数据揭示了一种细胞获取蛋白激酶的新模式，并确定了GSK3在精子成熟和顶体胞吐调节中的关键作用。
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