The 2004 Nobel Price in chemistry has been awarded to Aaron Ciechanover, Avram Hershko and Irvin Rose for the discovery of ATP-dependent, proteolytic ubiquitin system. The covalent binding of small protein ubiquitin to other proteins selectively targets such “ubiquitinated” proteins for degradation by the 26-S proteasome, a barrel-shaped multi-subunit protease complex. The eukaryotic ubiquitin system is highly substrate-specific and highly conserved at the same time, a versatility that allows it to modulate a variety of physiological and pathological responses ranging from cell cycle control to apoptosis. While ubiquitin research has grown into a major filed of cell biology, little is known about ubiquitin in the reproductive system.
The assertion that the sperm borne 26S proteasome takes part in the fertilization process from acrosomal exocytosis triggered by the egg coat glycoproteins to penetration of the vitelline coat and fusion with the egg plasma membrane (Mykles D., 1998, Int. Rev. Cytol. 184:157-289) been recently supported by new data from several laboratories around the world. Unexpectedly, a non-conventional, extracellular ubiquitin system has been uncovered (reviewed by Sakai et al., 2004, Int. J. Biochem. Cell Biol. 36:776-84) that participates in gametogenesis and gamete interactions during fertilization.
*Pines J. Nature 1994, 371:742-743.
Ubiquitin-proteasome Pathway during Spermatogenesis, Fertilization, and Preimplantation Embryo Development: Killing Three Birds with One Stone**
Dr. Sutovsky’s research has implicated the ubiquitin-proteasome pathway in the control of mitochondrial inheritance after mammalian fertilization and in the proteolytic mechanism for the sperm quality control during spermatogenesis. The central hypothesis of this research is that the maturation and function of human and animal spermatozoa is guided by the ubiquitin system in the testis, epididymis, and during/after fertilization. The components of the ubiquitin system in the spermatozoon are compartmentalized and developmentally regulated. The respective ubiquitin-dependent machineries regulate spatially and temporally separated events during the sperm cell life, such as sperm production in the testis, sperm maturation in the epididymis, penetration of the egg coat, zona pellucida during fertilization, targeted degradation of the paternal mitochondria, harboring paternal mitochondrial genes inside the fertilized egg.
The work on sperm production and maturation identified ubiquitin as an objective marker of sperm defects, leading to development of accurate male infertility tests for human diagnostic use and fertility assays for improving reproductive efficiency in farm animals (Sutovsky et al., 2001, J. Cell Sci. 114, 1665-1675; Sutovsky et al., 2001, Human Reprod. , 16, 250-258).
The description of the 26S-proteasome-assisted sperm-egg coat penetration could result in the development of a new, reversible, non-hormonal contraceptive (Sutovsky et al., 2004, Biol. Reprod. Reprod. 71, 1625–1637). Unique, sperm-specific cytoskeletal proteins may be responsible for anchoring of the 26S proteasome to sperm acrosomal membranes, matrix and subacrosomal cytoskeleton (Yu et al., 2006, Dev. Biol. 290:32-43).
The work on mitochondrial inheritance provided an explanation of how the fertilized egg destroys paternal mitochondrial genes (Sutovsky et al., 1999, Nature 402, 371-372: Sutovsky et al., 2003, Biol. Reprod., 68:1793-1800.). As a consequence of such selective sperm mitochondrion destruction, mitochondrial genes in humans and other mammals are inherited from the mother. This clonal inheritance pattern allows scientists to determine the lineage of human and animal evolution and to calibrate the evolutionary clock. Ubiquitin-proteasome-pathway is also important for early development after fertilization and for the remodeling of the donor cell after nuclear transfer/cloning of farm animals.
**Reviewed by Sutovsky, 2003, Micros. Res. Tech., 61: 88-102; Baska and Sutovsky, 2005. In: New impact on protein modifications in the regulation of reproductive system. Tokumoto, T, Ed., Research Signpost, Kerala.
Research on Sperm Perinuclear Theca (PT)***
Perinuclear theca (PT) is the protective coat of mammalian sperm head that is removed at fertilization. The PT harbors the sperm borne, oocyte-activating factors (SOAF), a yet uncharacterized complex responsible for triggering the signaling cascade of oocyte activation leading to the activation of embryo development and cleavage. During natural fertilization, the SOAF is transferred from the sperm PT to oocyte cytoplasm through the internalization and rapid solubilization of the sperm PT (Sutovsky et al., 1997, Dev. Biol. 188:75-84). After intracytoplasmic sperm injection (ICSI), a common treatment procedure for human infertility, oocyte activation occurs only in those oocytes in which the injected spermatozoon displays complete or partial solubilization of PT (Sutovsky et al., 1996, Human Reprod. 11:1703-1712). In some cases, the residual, subacrosomal layer of PT may persist on the surface of the sperm head after ISCI and may cause a delay or a complete arrest of embryo development. Improved ISCI procedure has been developed to prevent retention of sperm PT and increase fertilization rates after ICSI (Katayama et al., 2005, Reproduction 130:907-916). Sutovsky’s research on sperm PT is in collaboration with Dr. Richard Oko (Queen’s University, Kingston, ON) who has identified and characterized a novel sperm PT protein designated PT32/PAWP that is a candidate component of the SOAF involved in the induction of early development.
***Reviewed by Sutovsky et al., 2003, Microsc. Res. Tech., 61:362-378.
Insidermedicine In the Spotlight - Dr. Richard Oko
This research includes the development of the objective, automated assays and simple on-site test kits for an unbiased evaluation of semen quality in farm animals and the diagnosis of human male infertility. A patented procedure for rapidly and inexpensively assessing the fertility of genetically-superior male farm animals has already being developed. This test may eventually replace the current use of breeding large numbers of females to each male tested and then waiting for the pregnancy outcome — an approach that is very expensive and time consuming. The targeting of sperm ubiquitin-proteasome system for the purpose of developing an efficient non-hormonal contraceptive, and the improvement of assisted reproductive technologies such as ICSI and somatic cell nuclear transfer (SCNT) will be possible after deciphering the role of the ubiquitin system and sperm perinuclear theca in oocyte activation, pronuclear development, processing of the sperm accessory structures after ICSI and remodeling of the donor cell nucleus inside the recipient cytoplast after NT. Moreover, this work has broad implications for basic understanding of cell function. Sutovsky lab has uncovered several unconventional pathways employing ubiquitin-mediated protein recycling, during reproductive process, thereby adding to what was previously known about the vital cellular process.
****Reviewed by Sutovsky et al., 2003, Microsc. Res. Tech., 61:362-378; Prather et al., 2004, Exp. Biol. Med., 229: 1120-1126.