Sperm cannot fertilize an egg until sterols have been removed from the sperm plasma membrane by extracellular acceptors as part of the process of """"""""capacitation."""""""" Although the requirement for this re-ordering of lipids has been known for decades, several critical questions remain unanswered. How are lipids organized in the sperm plasma membrane? What molecules mediate sterol efflux? How is sterol efflux transduced into the functional changes that enable the sperm to fertilize? The health relevance of answering these questions include improving diagnostics for male infertility, improving sperm cryopreservation and handling in vitro, and helping design male contraceptives. These studies will advance our understanding of how a membrane's lipid microenvironment affects the activities of single proteins and signaling pathways. Such knowledge might also benefit conditions such as cardiac disease or diabetes in which the normal sterol metabolism of cells is perturbed. The recent discovery by the PI that the sperm plasma membrane is organized into sub-domains known as """"""""lipid rafts"""""""" presents an exciting link between sterol efflux and changes in sperm function. Lipid rafts are sub-domains of membrane enriched in sterols and sphingolipids (such as gangliosides), which both organize and regulate signaling pathways in somatic cells. New data from my laboratory are presented in this proposal which have led to the creation of the following model: 1) sperm possess at least two sub-types of lipid raft, one enriched in sterols and one enriched in the ganglioside, GM1, 2) in non-capacitated sperm, these rafts are strictly segregated by interactions with underlying cytoskeletal proteins, 3) sterol efflux during capacitation causes dissolution of sterol-enriched rafts and re-distribution of rafts enriched in GM1, and 4) these alterations in the lipid microenvironment cause changes in the activities of membrane proteins, enabling the sperm to fertilize. The model will be tested by characterizing the lipid and protein content of the lipid rafts (Aim 1). Both targeted (e.g. immunoblots) and proteomic (e.g. two dimensional gel electrophoresis and protein sequencing) experimental approaches will determine which cytoskeletal proteins interact with lipid rafts, and how they regulate dynamic movements of rafts (Aim 2). Studying the ability and kinetics of different sterol acceptors will help deduce the nature of the protein(s) mediating efflux (Aim 3). The membrane lipid content of a heterologous expression system as well as of sperm will be altered to test the effect of sterols and gangliosides on the activity of a potassium channel and a sperm's ability to undergo acrosomal exocytosis (Aim 4).

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD045664-04
Application #
7150004
Study Section
Reproductive Biology Study Section (REB)
Program Officer
Rankin, Tracy L
Project Start
2004-01-16
Project End
2008-11-30
Budget Start
2006-12-01
Budget End
2007-11-30
Support Year
4
Fiscal Year
2007
Total Cost
$269,663
Indirect Cost
Name
Cornell University
Department
Other Basic Sciences
Type
Schools of Veterinary Medicine
DUNS #
872612445
City
Ithaca
State
NY
Country
United States
Zip Code
14850
Cohen, Roy; Buttke, Danielle E; Asano, Atsushi et al. (2014) Lipid modulation of calcium flux through CaV2.3 regulates acrosome exocytosis and fertilization. Dev Cell 28:310-21
Asano, Atsushi; Nelson-Harrington, Jacquelyn L; Travis, Alexander J (2013) Phospholipase B is activated in response to sterol removal and stimulates acrosome exocytosis in murine sperm. J Biol Chem 288:28104-15
Asano, Atsushi; Nelson, Jacquelyn L; Zhang, Sheng et al. (2010) Characterization of the proteomes associating with three distinct membrane raft sub-types in murine sperm. Proteomics 10:3494-505
Selvaraj, Vimal; Asano, Atsushi; Page, Jennifer L et al. (2010) Mice lacking FABP9/PERF15 develop sperm head abnormalities but are fertile. Dev Biol 348:177-89
Travis, A J; Kim, Y; Meyers-Wallen, V (2009) Development of new stem cell-based technologies for carnivore reproduction research. Reprod Domest Anim 44 Suppl 2:22-8
Asano, Atsushi; Selvaraj, Vimal; Buttke, Danielle E et al. (2009) Biochemical characterization of membrane fractions in murine sperm: identification of three distinct sub-types of membrane rafts. J Cell Physiol 218:537-48
Mukai, Chinatsu; Bergkvist, Magnus; Nelson, Jacquelyn L et al. (2009) Sequential reactions of surface- tethered glycolytic enzymes. Chem Biol 16:1013-20
Selvaraj, Vimal; Asano, Atsushi; Buttke, Danielle E et al. (2009) Mechanisms underlying the micron-scale segregation of sterols and GM1 in live mammalian sperm. J Cell Physiol 218:522-36
Kim, Yeunhee; Turner, Danielle; Nelson, Jacquelyn et al. (2008) Production of donor-derived sperm after spermatogonial stem cell transplantation in the dog. Reproduction 136:823-31
Simpson, Ian A; Dwyer, Donard; Malide, Daniela et al. (2008) The facilitative glucose transporter GLUT3: 20 years of distinction. Am J Physiol Endocrinol Metab 295:E242-53

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