|Rafael A. Fissore
Associate Professor of Veterinary and Animal Sciences, University of Massachusetts
Ph.D.: University of Massachusetts
Ca2+ Release Mechanisms in Mammalian Oocytes during Maturation and Fertilization
The sperm during fertilization initiates in mammalian oocytes a series of [Ca2+]i oscillations that are critical for activation and initiation of development. Two hypotheses have been suggested to explain the signaling mechanism(s) utilized by the sperm to trigger Ca2+ release, the receptor hypothesis and the fusion hypothesis. The fusion hypothesis predicts that a sperm cytosolic protein is released into the oocyte upon fusion. In accordance with this theory, extracts from sperm induce, when injected into oocytes, [Ca2+]i oscillations similar to those observed during fertilization. Our extracts, from porcine sperm, have the ability to induce high frequency Ca2+ release in mouse oocytes. Partial characterization of the active fraction revealed a pI of 6.7-7.0, a relative molecular weight of 29-68 kDa, and absence of gpi/oscillin, a protein previously thought to represent the active component of mammalian sperm. Future studies will establish the molecular nature of the active compound(s) and its possible site of action in mammalian eggs.
In another project, we are characterizing the role of mos and MAP kinase in the initiation, and progression, of meiosis in mammalian oocytes. We are also interested in investigating the role of Ca2+ in the activation of maturation promoting factor (MPF) in the same stage of meiosis. For these experiments, mRNA’s coding for different proteins are injected into oocytes and the effects on kinase activity and meiosis progression is assessed by in vitro kinase assays and by Hoescht staining. Ca2+ activity is assessed by microfluorometry. Other general molecular and cellular biology techniques are used in the laboratory.
Gordo A.C., Rodrigues P., Kurokawa M., Jellerette T., Exley G., Warner C., and R.A. Fissore (2002). [Ca 2+ ]i Oscillations Signal Apoptosis Rather than Activation in In vitro Aged Mouse Eggs. Biology of Reproduction 66, 1828-1837.
Gordo A.C., Kurokawa M., Wu H., and R.A. Fissore (2002). Modifications of the Ca 2+ release mechanisms of mouse oocytes by fertilization and by sperm factor. Molecular Human Reproduction 8, 619-629.
Smyth J.T., Abbott A.L., Lee B., Sienaert I., Nadif Kasri N., De Smedt H., Ducibella T., Missiaen L., Parys J.B., and R.A. Fissore (2002). KN-93 inhibits the type 1 inositol trisphosphate receptor independently of Ca 2+ /calmodulin protein kinase II inhibition. J. Biol. Chem. In Press.
Knott, J. G., K. Poothapillai, H. Wu, C. L. He, R. A. Fissore and J. M. Robl (2002). "Porcine sperm factor supports activation and development of bovine nuclear transfer embryos." Biol Reprod 66(4): 1095-103.
Abbott, A. L., R. A. Fissore and T. Ducibella (2001). "Identification of a translocation deficiency in cortical granule secretion in preovulatory mouse oocytes." Biol Reprod 65(6): 1640-7.
Fukami, K., K. Nakao, T. Inoue, Y. Kataoka, M. Kurokawa, R. A. Fissore, K. Nakamura, M. Katsuki, K. Mikoshiba, N. Yoshida and T. Takenawa (2001). "Requirement of phospholipase Cdelta4 for the zona pellucida-induced acrosome reaction." Science 292(5518): 920-3.
Lee, S. C., R. A. Fissore and R. Nuccitelli (2001). "Sperm factor initiates capacitance and conductance changes in mouse eggs that are more similar to fertilization than IP(3)- or Ca(2+)-induced changes." Dev Biol 232(1): 127-48.
Salamone, D. F., P. Damiani, R. A. Fissore, J. M. Robl and R. T. Duby (2001). "Biochemical and developmental evidence that ooplasmic maturation of prepubertal bovine oocytes is compromised." Biol Reprod 64(6): 1761-8.
Wu, H., J. Smyth, V. Luzzi, K. Fukami, T. Takenawa, S. L. Black, N. L. Allbritton and R. A. Fissore (2001). "Sperm factor induces intracellular free calcium oscillations by stimulating the phosphoinositide pathway." Biol Reprod 64(5): 1338-49.
Gordo, A. C., C. L. He, S. Smith and R. A. Fissore (2001). "Mitogen activated protein kinase plays a significant role in metaphase II arrest, spindle morphology, and maintenance of maturation promoting factor activity in bovine oocytes." Mol Reprod Dev 59(1): 106-14.
Stricker SA, Swann K, Jones KT, Fissore RA. (2000). Injections of Porcine sperm extracts trigger fertilization-like Ca2+ oscillations in oocytes of a marine worm. Experimental Cell Research. 257: 341-347.
Jellerette T, He CL, Wu H, Parys JB, Fissore RA. (2000). Down-regulation of the inositol 1,4,5,-trisphosphate receptor in mouse eggs following fertilization or parthenogenetic activation. Developmental Biology 223, 238-250.
Gordo AC, Wu H, He CL, Fissore RA. (2000). Injection of sperm cytosolic factor into mouse Metaphase II oocytes induces different developmental outcomes according to the frequency of [Ca2+]i oscillations and oocyte age. Biology of Reproduction 62: 1370-1379.