Isoform-specific requirement for GSK3α in sperm for male fertility†

Abstract

Glycogen synthase kinase 3 (GSK3) is a highly conserved protein kinase regulating key cellular functions. Its two isoforms, GSK3α and GSK3β, are encoded by distinct genes. In most tissues the two isoforms are functionally interchangeable, except in the developing embryo where GSK3β is essential. One functional allele of either of the two isoforms is sufficient to maintain normal tissue functions. Both GSK3 isoforms, present in sperm from several species including human, are suggested to play a role in epididymal initiation of sperm motility. Using genetic approaches, we have tested requirement for each of the two GSK3 isoforms in testis and sperm. Both GSK3 isoforms are expressed at high levels during the onset of spermatogenesis. Conditional knockout of GSK3α, but not GSK3β, in developing testicular germ cells in mice results in male infertility. Mice lacking one allele each of GSK3α and GSK3β are fertile. Despite overlapping expression and localization in differentiating spermatids, GSK3β does not substitute for GSK3α. Loss of GSK3α impairs sperm hexokinase activity resulting in low ATP levels. Net adenine nucleotide levels in caudal sperm lacking GSK3α resemble immature caput epididymal sperm. Changes in the association of the protein phosphatase PP1γ2 with its protein interactors occurring during epididymal sperm maturation is impaired in sperm lacking GSK3α. The isoform-specific requirement for GSK3α is likely due to its specific binding partners in the sperm principal piece. Testis and sperm are unique in their specific requirement of GSK3α for normal function and male fertility.

Roles of glycogen synthase kinase 3 alpha and calcineurin in regulating the ability of sperm to fertilize eggs

“…20,21,23 The sperm phenotypes of the KO are similar. The two signaling enzymes, GSK3α and PPP3R2, are essential for male fertility: deletion of either corresponding genes results in male infertility.…”

“…Indeed, it has been reported that the luminal fluid of caput has threefold higher calcium concentration than the caudal region of the epididymis. 20 The next part of our study was examination of the relationship between sperm calcineurin and GSK3. [59][60][61] A key conclusion of our study is that high calcineurin activity in caput sperm followed by a decline in its activity in caudal sperm is a key physiological requirement for epididymal sperm maturation.…”

“…20 Antigen retrieval was performed using 1X Antigen Retrieval Citra Solution (BioGenex, San Ramon, CA, USA). The fixed testes were transferred to 75% ethanol and dehydrated, permeabilized, and embedded in paraffin using a Shandon Tissue Processor (Thermo Electron Corp. Waltham, MA, USA).…”

“…12,14 These findings lead to a model wherein cAMP action through PKA is the central tenet of the mechanistic scheme. 20,21 Thus, GSK3α is essential in testis and sperm. 15 We showed that a serine/threonine phosphatase (PP1) has a key role in sperm maturation and regulation of motility.…”

“…These models however fall short because they do not incorporate protein phosphatases, which are known to be initiators, modifiers, and terminators of signaling responses in cells. 20 Interestingly, the activity of GSK3 declines during epididymal sperm maturation: that is, catalytically active GSK3 is required in maturing epididymal sperm. 16,17 The protein phosphatase isoform PP1γ2, present only in mammals, is regulated by its binding proteins-PPP1R2, PPP1R7, and PPP1R11.…”