Characterization of a novel role for the dynamin mechanoenzymes in the regulation of human sperm acrosomal exocytosis.

Show simple item record Zhou, Wei en Anderson, Amanda L en Turner, Adrian en De Iuliis, Geoffry N en McCluskey, Adam en McLaughlin, Eileen en Nixon, Brett en 2018-11-16T02:53:46Z en 2017-10 en
dc.identifier.citation Molecular human reproduction 23(10):657-673 Oct 2017 en
dc.identifier.issn 1360-9947 en
dc.identifier.uri en
dc.description.abstract STUDY QUESTION:Does dynamin regulate human sperm acrosomal exocytosis? SUMMARY ANSWER:Our studies of dynamin localization and function have implicated this family of mechanoenzymes in the regulation of progesterone-induced acrosomal exocytosis in human spermatozoa. WHAT IS KNOWN ALREADY:Completion of an acrosome reaction is a prerequisite for successful fertilization in all studied mammalian species. It follows that failure to complete this unique exocytotic event represents a common aetiology in the defective spermatozoa of male infertility patients that have failed IVF in a clinical setting. Recent studies have implicated the dynamin family of mechanoenzymes as important regulators of the acrosome reaction in murine spermatozoa. The biological basis of this activity appears to rest with the ability of dynamin to polymerize around newly formed membrane vesicles and subsequently regulate the rate of fusion pore expansion. To date, however, the dynamin family of GTPases have not been studied in the spermatozoa of non-rodent species. Here, we have sought to examine the presence and functional significance of dynamin in human spermatozoa. STUDY DESIGN, SIZE, DURATION:Dynamin expression was characterized in the testis and spermatozoa of several healthy normozoospermic individuals. In addition, we assessed the influence of selective dynamin inhibition on the competence of human spermatozoa to undergo a progesterone-induced acrosome reaction. A minimum of five biological and technical replicates were performed to investigate both inter- and intra-donor variability in dynamin expression and establish statistical significance in terms of the impact of dynamin inhibition. PARTICIPANTS/MATERIALS, SETTING, METHODS:The expression and the localization of dynamin in the human testis, epididymis and mature spermatozoa were determined through the application of immunofluorescence, immunoblotting and/or electron microscopy. Human semen samples were fractionated via density gradient centrifugation and the resultant populations of good and poor quality spermatozoa were induced to capacitate and acrosome react in the presence or absence of selective dynamin inhibitors. The acrosome integrity of live spermatozoa was subsequently assessed via the use of fluorescently conjugated Arachis hypogea lectin (PNA). The influence of dynamin phosphorylation and the regulatory kinase(s) responsible for this modification in human spermatozoa were also assessed via the use of in situ proximity ligation assays and pharmacological inhibition. In all experiments, ≥100 spermatozoa were assessed/treatment group and all graphical data are presented as the mean values ± SEM, with statistical significance being determined by ANOVA. MAIN RESULTS AND THE ROLE OF CHANCE:Dynamin 1 (DNM1) and DNM2, but not DNM3, were specifically localized to the acrosomal region of the head of human spermatozoa, an ideal position from which to regulate acrosomal exocytosis. In keeping with this notion, pharmacological inhibition of DNM1 and DNM2 was able to significantly suppress the rates of acrosomal exocytosis stimulated by progesterone. Furthermore, our comparison of dynamin expression in good and poor quality spermatozoa recovered from the same ejaculate, revealed a significant reduction in the amount of DNM2 in the latter subpopulation of cells. In contrast, DNM1 was detected at equivalent levels in both subpopulations of spermatozoa. Such findings are of potential significance given that the poor quality spermatozoa proved refractory to the induction of a progesterone stimulated acrosome reaction. In seeking to identify the regulatory influence of progesterone on DNM2 function, we were able to establish that the protein is a substrate for CDK1-dependent phosphorylation. The functional significance of DNM2 phosphorylation was illustrated by the fact that pharmacological inhibition of CDK1 elicited a concomitant suppression of both DNM2-Ser764 phosphorylation and the overall rates of progesterone-induced acrosomal exocytosis. LARGE SCALE DATA:N/A. LIMITATIONS REASONS FOR CAUTION:This was an in vitro study performed mainly on ejaculated human spermatozoa. This experimental paradigm necessarily eliminates the physiological contributions of the female reproductive tract that would normally support capacitation and acrosomal responsiveness. WIDER IMPLICATIONS OF THE FINDINGS:This study identifies a novel causative link between dynamin activity and the ability of human spermatozoa to complete a progesterone-induced acrosome reaction. Such findings encourage a more detailed analysis of the contribution of dynamin dysregulation as an underlying aetiology in infertile males whose spermatozoa are unable to penetrate the zona pellucida. STUDY FUNDING/COMPETING INTEREST(S):This research was supported by a National Health and Medical Research Council of Australia Project Grant (APP1103176) awarded to B.N. and E.A.M. The authors report no conflict of interest. en
dc.format.medium Print en
dc.language eng en
dc.relation.ispartofseries Molecular human reproduction en
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. en
dc.rights This is a pre-copyedited, author-produced version of an article accepted for publication in Molecular Human Reproduction following peer review. The version of record is available online at: en
dc.rights.uri en
dc.rights.uri en
dc.subject Testis en
dc.subject Epididymis en
dc.subject Spermatozoa en
dc.subject Brain en
dc.subject Animals en
dc.subject Humans en
dc.subject Mice en
dc.subject Hydrazones en
dc.subject Naphthols en
dc.subject Progesterone en
dc.subject Dynamins en
dc.subject Dynamin I en
dc.subject CDC2 Protein Kinase en
dc.subject Protein Isoforms en
dc.subject Mechanotransduction, Cellular en
dc.subject Exocytosis en
dc.subject Gene Expression Regulation en
dc.subject Phosphorylation en
dc.subject Acrosome Reaction en
dc.subject Male en
dc.title Characterization of a novel role for the dynamin mechanoenzymes in the regulation of human sperm acrosomal exocytosis. en
dc.type Journal Article en
dc.identifier.doi 10.1093/molehr/gax044 en
pubs.issue 10 en
pubs.begin-page 657 en
pubs.volume 23 en
dc.rights.holder Copyright: The authors en
dc.identifier.pmid 29044420 en
pubs.end-page 673 en
pubs.publication-status Published en
dc.rights.accessrights en
pubs.subtype Journal Article en
pubs.elements-id 698732 en
dc.identifier.eissn 1460-2407 en
pubs.record-created-at-source-date 2017-10-19 en
pubs.dimensions-id 29044420 en

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