- Title
- Functional characterisation of dynamin in mouse spermatozoa
- Creator
- Reid, Andrew Timothy
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2015
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- World population growth has been increasing exponentially since the 1950’s, highlighting the inadequacies of current methods of contraception and the urgent need for new contraceptive measurements. Conversely, although the major driver of this population growth is the large number of unplanned pregnancies in developing countries, male factor infertility is on the rise within the developed world where it is now estimated to afflict 1 in every 20 males. To address these paradoxical problems we need a far better understanding of the mechanisms behind fertilization, with particular focus on the spermatozoon. Such knowledge would enable us to identify new contraceptive targets as well improve diagnosis of the underlying causes of infertility. Two key functions that the spermatozoon must fulfil in order to fertilize in vivo are spermzona pellucida (ZP) binding and acrosomal exocytosis. In order for spermatozoa to obtain the ability to both bind the ZP and undergo a stimulus induced acrosome reaction they must undergo three distinct phases of maturation, namely spermatogenesis, epididymal maturation and capacitation. In the studies described in this thesis we have sought to investigate the role of the large membrane manipulator, dynamin, during sperm maturation as well as investigate its role during key phases of the fertilization cascade. We have found that dynamin is present within the developing acrosome of immature gametes within the testis and is retained in this domain throughout epididymal maturation and capacitation. Via pharmacological inhibition of dynamin we have demonstrated that the GTPase is responsible for regulation of the progesterone induced acrosome reaction and this directly influences the rates of in vitro fertilization. Furthermore, upon receiving this stimulus to acrosome react, dynamin becomes phosphorylated at key serine residues. Subsequent to these findings we investigated these phosphorylation events further and identified the kinase responsible for the key phosphorylation event on serine 774 of dynamin 1 as glycogen synthase kinase 3 (GSK3). We have demonstrated via pharmacological inhibition of GSK3 that its activity is pivotal in controlling the onset of acrosomal exocytosis. Remarkably, via immunoelectron microscopy we have shown the delivery of this dynamin-phosphorylating kinase to sperm within the distal corpus epididymis via novel granular structures referred to as ‘dense bodies’. Finally we have determined that dynamin 1 and dynamin 2, are positioned to play important regulatory roles within the principal cells as well as the lumen of the epididymis. We have provided evidence that these proteins play essential roles in maintaining the composition of the intraluminal milieu across the caput, corpus and cauda epididymis, and thus influence the maturation of spermatozoa. Taken together, these data have demonstrated a previously unappreciated role for dynamin during pivotal stages of spermatogenesis as well as during the epididymal maturation of spermatozoa. Furthermore, it is clear that dynamin function is essential for fertilization via its ability to regulate the acrosome reaction. Such findings add considerably to our understanding of the molecular mechanisms that underpin the production of functionally competent spermatozoa and will likely help to guide future research into contraceptive development as well as infertility studies.
- Subject
- spermatozoa; Dynamin; acrosome reaction
- Identifier
- http://hdl.handle.net/1959.13/1310603
- Identifier
- uon:22056
- Rights
- Copyright 2015 Andrew Timothy Reid
- Language
- eng
- Full Text
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| View Details Download | ATTACHMENT02 | Thesis | 8 MB | Adobe Acrobat PDF | View Details Download |