Fertilisation and Contraception
Sperm Oocyte Interaction
Formation of Sperm Zona Receptor Complex
Mammalian sperm-egg interaction is arguably one of the most remarkable processes in biological science. This exquisitely specific cell recognition event depends upon a complex cascade of interactions between free-swimming sperm and ovulated eggs. Elucidating the nature of these interactions has been the subject of intense investigation by many laboratories. Although this has led to extensive characterization of the respective gametes, such studies have failed to elucidate the molecular basis of this event. In our considered judgement this lack of success stems from the incorrect assumption that the sperm receptor is a single molecular entity that is constitutively expressed on the cell surface.
In contrast, recent research from our laboratory has provided support for a novel hypothesis that sperm-egg interaction is mediated by a multimeric sperm receptor complex. Furthermore, we have compelling evidence that this complex is assembled on the sperm surface through the concerted action of a family of molecular chaperone proteins that reside within specialised membrane domains, known as lipid rafts. In this project we aim to confirm the validity of this model, establish the molecular composition of the sperm surface receptor complexes and characterise the downstream signalling cascades that culminate in fertilisation
Oolemma sperm receptors
Post-coitus, sperm encounter the ovulated oocyte within the fallopian tube, which serves as the site of fertilisation. By the time sperm reach the site of fertilisation they have undergone the prerequisite series of capacitation-associated changes necessary for ultimately achieving fertilisation. Upon penetrating the Zona pellucida, sperm must then recognise and bind to the oocyte plasma membrane. Together, the molecular machinery present in both cell membranes must then interact in a precise manner necessary to facilitate the energetically costly event that is the merger of sperm and egg membranes.
This research aims to identify and characterise egg surface proteins implicated in sperm-egg interaction, by identifying the important GPI-anchored protein(s) as well characterising the role of the tetraspanins and their interacting web partners. In vitro binding and fusion experiments will be used as functional bioassays and mass spectrometry (MS)-based proteomics and bioinformatics-based analyses will be employed to compile oocyte protein databases and to identify candidate proteins responsible for mediating sperm-egg interaction, such as newly identified candidate GPI-anchored proteins.
Spermicides and microbicides
There is an urgent clinical need to research novel methods of fertility control that are also protective against sexually transmitted diseases (STDs) such as the human immunodeficiency virus (HIV) or Chlamydia. The most obvious way to generate such a dual-purpose contraceptive method would be to develop safe, effective spermicides that were also active against a wide range of pathogenic organisms. The currently available formulations such as nonoxynol-9, gramicidin and benzalkonium chloride are effective spermicides but are toxic to the vaginal epithelium and do not provide protection against STDs. Over 60 agents are in clinical trials as potentially safer topical spermicides and/or microbicides. Compounds that have reached this stage of development include acid buffers, detergents, dendrimers, non-nucleoside reverse transcriptase inhibitors and anionic polymers. In addition, a number of potential spermicides/microbicides are the subject of preclinical investigation, including beta-cyclodextrin, cyanovirin, porphyrins, cyclotriazadisulfonamides, dermaseptins, short-interfering RNA (siRNA) and HIV antibodies. This project aims using to characterise the biological activities and mode of action of newly developed target specific compounds as novel dual spermicides/microbicides.
Environmental Impacts on Female Fertility
All female mammalian ovaries contain a limited supply of primordial follicles which are present from birth. Recently it has become apparent that xenobiotics, such as organochlorine pesticides, polychlorinated biphenyls, dioxins, alkyl phenolic chemicals, phthalates and synthetic oestrogens are capable of interfering with normal female reproductive function in both humans and animals. Some xenobiotics that are prevalent in the environment including 4-vinylcyclohexane and benzo[a]pyrene, have been shown to target primordial follicles and trigger atretic oocyte depletion of the ovary leading to premature menopause. Our recent studies of the effects of xenobiotic exposure in the ovary have demonstrated that environmental agents can cause significant primordial follicle loss and oocyte damage through oxidative stress. Our proposed model is based on preliminary findings which indicate that xenobiotic exposure has direct consequences on ovarian function.
The aims of this project are: To elucidate the action of xenobiotics in the developing ovary and in particular the role of the Phase I enzymes (Cytochrome p450's) in the generation of reactive oxygen species. To quantify the role of xenobiotic derived oxidative stress on mitochondrial function, plasma membrane fluidity and oocyte dysfunction. To characterize the ovarian follicle signalling pathways activated by xenobiotics and their role in oocyte growth, follicle differentiation and survival. Insight into these processes will illuminate the origins of primordial follicle loss and oocyte dysfunction leading to subsequent ovarian failure and infertility in human females.
What Makes a Good Egg?
There is major interest in the molecular mechanisms regulating the maintenance and development of oocytes in the mammalian ovary. Exhaustion of the supply of oocytes results in menopause. The deterioration of oocyte quality with age has significant impacts on female fertility, meiotic defects in the embryo, particularly trisomy 21, and long term susceptibility to conditions such as cardiac disease, diabetes and obesity. Crucial to ovarian function are RNA binding proteins, which control post-transcriptional regulation of mRNAs coding for proteins essential for germ cell and follicle development. We have localised Musashi 1 and Musashi 2, RNA binding proteins with known roles in stem cell specification and meiotic segregation, to the mouse oocyte.
This project aims to characterise the expression of Musashi 1 and 2 throughout oogenesis and folliculogenesis and to identify the mRNA target(s) of Musashi 1 and 2 for translational repression in the oocyte, delivering possible therapeutic options for improving human oocyte health.
Identification and validation of targets for fertility regulation
There have been no significant advances in planned fertility regulation since the introduction of the oral contraceptive pill in the 1960s. The purpose of this project is to use our advanced proteomics platform to identify targets for male and female fertility regulation on the basis of their specificity, functional significance and druggability. The current focus is on the identification of phosphoproteins that are critical to the process of fertilization and then through a knowledge of the kinases and phosphatases that regulate their phosphorylation statu,s to select contraceptive targets that can then be validated by functional genomics in vivo and in vitro.