Skip to main content Accessibility help
×
Home

Calcineurin role in porcine oocyte activation

  • L. Tůmová (a1), E. Chmelíková (a1), T. Žalmanová (a1), V. Kučerová-Chrpová (a1), R. Romar (a2), M. Dvořáková (a1), K. Hošková (a1) and J. Petr (a3)...

Abstract

Calcineurin is required for oocyte exit from meiotic block in metaphase II (MII) stage in invertebrates and also in lower vertebrates. However, the role of calcineurin in mammalian oocyte activation is still unclear. The aim of this study was to determine whether calcineurin is involved in the processes regulating porcine oocyte activation. Indirect immunofluorescence demonstrated localization of both calcineurin subunits, CnA and CnB, especially in the cortex area of MII oocytes, in vitro fertilized and also parthenogenetically activated oocytes. After activation, the fluorescence intensity of the protein in the cortex area of oocytes remains unchanged; the protein calcineurin in the cytoplasm was recorded mainly around the pronuclei. Treatment of matured oocytes with calcineurin inhibitors, cyclosporin A (CsA) and hymenistatin I (HS-I), followed by activation with calcium ionophore A23187, significantly decreased the rate of activated oocytes compared to oocytes that were treated only with calcium ionophore (Ca-Io), (CsA+Ca-Io 25.0% v. Ca-Io 83.3%; HS-I+Ca-Io 32.5% v. Ca-Io 85.0%). Compared to the control, CsA treatment of matured oocytes followed by activation with Ca-Io did not affect the activity level of metaphase-promoting factor (MPF) and mitogen-activated protein kinase (MAPK) in activated oocytes evaluated by kinase activity assay. Simultaneous staining of calcineurin and cortical granule content in matured oocytes showed that calcineurin distributed in the cortical area of the oocyte has not been colocalized with cortical granules content. On the other hand, the calcineurin inhibition before parthenogenetic activation leads to a reduction of the cortical reaction level compared to oocytes that were not treated with CsA (complete exocytosis: CsA+Ca-Io 2.6% v. Ca-Io 83.9%; sum of cortical granule brightness: CsA + Ca-Io 0.69 v. Ca-Io 0.15). Our results showed that calcineurin is involved in the process of pig oocyte activation and cortical granule exocytosis; however this regulation seems to be MPF and MAPK independent.

Copyright

Corresponding author

E-mail: chmelikova@af.czu.cz

References

Hide All
Abbott, AL and Ducibella, T 2001. Calcium and the control of mammalian cortical granule exocytosis. Frontiers in Bioscience 6, D792D806.
Bleil, JD, Beall, CF and Wassarman, PM 1981. Mammalian sperm-egg interaction – fertilization of mouse eggs triggers modification of the major zona pellucida glycoprotein, ZP2. Developmental Biology 86, 189197.
Chantler, PD 1985. Calcium-dependent association of a protein complex with the lymphocyte plasma-membrane – probable identity with calmodulin calcineurin. Journal of Cell Biology 101, 207216.
Depreux, FFS, Scheffler, JM, Grant, AL, Bidwell, CA and Gerrard, DE 2010. Molecular cloning and characterization of porcine calcineurin-alpha subunit expression in skeletal muscle. Journal of Animal Science 88, 562571.
Ducibella, T 1996. The cortical reaction and development of activation competence in mammalian oocytes. Human Reproduction Update 2, 2942.
Dupont, G 1998. Link between fertilization-induced Ca2+oscillations and relief from metaphase II arrest in mammalian eggs: a model based on calmodulin-dependent kinase II activation. Biophysical Chemistry 72, 153167.
Feine, O, Hukasova, E, Bruinsma, W, Freire, R, Fainsod, A, Gannon, J, Mahbubani, HM, Lindqvist, A and Brandeis, M 2014. Phosphorylation-mediated stabilization of Bora in mitosis coordinates Plx1/Plk1 and Cdk1 oscillations. Cell Cycle 13, 17271736.
Horner, VL and Wolfner, MF 2008. Transitioning from egg to embryo: triggers and mechanisms of egg activation. Developmental Dynamics 237, 527544.
Jilek, F, Huttelova, R, Petr, J, Holubova, M and Rozinek, J 2001. Activation of pig oocytes using calcium ionophore: effect of the protein kinase inhibitor 6-dimethyl aminopurine. Reproduction in Domestic Animals 36, 139145.
Kheilová, K, Petr, J, Žalmanová, T, Kučerová-Chrpová, V and Řehák, D 2015. Src family kinases are involved in the meiotic maturation of porcine oocytes. Reproduction, Fertility and Development 27, 10971105.
Klee, CB, Draetta, GF and Hubbard, MJ 1988. Calcineurin. Advances in Enzymology and Related Areas of Molecular Biology 61, 149200.
Krauchunas, AR, Sackton, KL and Wolfner, MF 2013. Phospho-regulation pathways during egg activation in Drosophila melanogaster . Genetics 195, 171180+.
Kubelka, M, Motlik, J, Schultz, RM and Pavlok, A 2000. Butyrolactone I reversibly inhibits meiotic maturation of bovine oocytes, without influencing chromosome condensation activity. Biology of Reproduction 62, 292302.
Lee, J, Miyano, T and Moor, RM 2000. Localisation of phosphorylated MAP kinase during the transition from meiosis I to meiosis II in pig oocytes. Zygote 8, 119125.
Liu, J, Farmer, JD, Lane, WS, Friedman, J, Weissman, I and Schreiber, SL 1991. Calcineurin is a common target of cyclophilin-cyclosporine-A and FKBP-FK506 complexes. Cell 66, 807815.
Longo, FJ and Chen, DY 1985. Development of cortical polarity in mouse eggs – involvement of the meiotic apparatus. Developmental Biology 107, 382394.
Lorca, T, Cruzalegui, FH, Fesquet, D, Cavadore, JC, Mery, J, Means, A and Doree, M 1993. Calmodulin-dependent protein kinase-II mediates inactivation of MPF and CSF upon fertilization of Xenopus eggs. Nature 366, 270273.
Manders, EMM, Verbeek, FJ and Aten, JA 1993. Measurement of co-localization of objects in dual-color confocal images. Journal of Microscopy 169, 375382.
Miyano, T, Dai, YF, Lee, J, Kano, K and Moor, RM 2000. Degradation of pig cyclin B1 molecules precedes MAP kinase dephosphorylation during fertilisation of the oocytes. Zygote 8, 153158.
Miyazaki, S, Yuzaki, M, Nakada, K, Shirakawa, H, Nakanishi, S, Nakade, S and Mikoshiba, K 1992. Block of Ca2+wave and Ca2+oscillation by antibody to the inositol 1,4,5-trisphosphate receptor in fertilized hamster eggs. Science 257, 251255.
Mochida, S and Hunt, T 2007. Calcineurin is required to release Xenopus egg extracts from meiotic M phase. Nature 449, 336340+.
Moos, J, Visconti, PE, Moore, GD, Schultz, RM and Kopf, GS 1995. Potential role of mitogen-activated protein-kinase in pronuclear envelope assembly and disassembly following fertilization of mouse eggs. Biology of Reproduction 53, 692699.
Nishiyama, T, Yoshizaki, N, Kishimoto, T and Ohsumi, K 2007. Transient activation of calcineurin is essential to initiate embryonic development in Xenopus laevis. Nature 449, 341345.
Petr, J, Chmelikova, E, Zalmanova, T, Tumova, L, Kheilova, K, Kucerova-Chrpova, V and Jilek, F 2013. Pyrethroids cypermethrin, deltamethrin and fenvalerate have different effects on in vitro maturation of pig oocytes at different stages of growth. Animal 7, 134142.
Pursel, VG and Johnson, LA 1975. Freezing of boar spermatozoa – fertilizing-capacity with concentrated semen and a new thawing procedure. Journal of Animal Science 40, 99102.
Romar, R, Coy, P, Gadea, J and Rath, D 2005. Effect of oviductal and cumulus cells on zona pellucida and cortical granules of porcine oocytes fertilized in vitro with epididymal spermatozoa. Animal Reproduction Science 85, 287300.
Rusnak, F and Mertz, P 2000. Calcineurin: form and function. Physiological Reviews 80, 14831521.
Saavedra, MD, Mondéjar, I, Coy, P, Betancourt, M, González-Márquez, H, Jiménez-Movilla, M, Avilés, M and Romar, R 2014. Calreticulin from suboolemmal vesicles affects membrane regulation of polyspermy. Reproduction 147, 369378.
Saunders, CM, Larman, MG, Parrington, J, Cox, LJ, Royse, J, Blayney, LM, Swann, K and Lai, FA 2002. PLC zeta: a sperm-specific trigger of Ca(2+) oscillations in eggs and embryo development. Development 129, 35333544.
Schatten, G 1994. The centrosome and its mode of inheritance – the reduction of the centrosome during gametogenesis and its restoration during fertilization. Developmental Biology 165, 299335.
Silverman-Gavrila, LB and Charlton, MP 2009. Calcineurin and cytoskeleton in low-frequency depression. Journal of Neurochemistry 109, 716732.
Takeo, S, Hawley, RS and Aigaki, T 2010. Calcineurin and its regulation by Sra/RCAN is required for completion of meiosis in Drosophila . Developmental Biology 344, 957967.
Takeo, S, Tsuda, M, Akahori, S, Matsuo, T and Aigaki, T 2006. The calcineurin regulator Sra plays an essential role in female meiosis in Drosophila . Current Biology 16, 14351440.
Tůmová, L, Petr, J, Žalmanová, T, Chmelíková, E, Kott, T, Tichovská, H, Kučerová-Chrpová, V, Hošková, K and Jílek, F 2013. Calcineurin expression and localisation during porcine oocyte growth and meiotic maturation. Animal Reproduction Science 141, 154163.
Yamamuro, T, Kano, K and Naito, K 2008. Functions of FZR1 and CDC20, activators of the anaphase-promoting complex during meiotic maturation of swine oocytes. Biology of Reproduction 79, 12021209.

Keywords

Type Description Title
WORD
Supplementary materials

Tůmová supplementary material
Table S1 and Figures S1-S4

 Word (3.1 MB)
3.1 MB

Calcineurin role in porcine oocyte activation

  • L. Tůmová (a1), E. Chmelíková (a1), T. Žalmanová (a1), V. Kučerová-Chrpová (a1), R. Romar (a2), M. Dvořáková (a1), K. Hošková (a1) and J. Petr (a3)...

Metrics

Altmetric attention score

Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed