Parthenogenetic activation of bovine oocytes using bovine and murine phospholipase C zeta.
BACKGROUND
During natural fertilization, sperm fusion with the oocyte induces long lasting intracellular calcium oscillations which in turn are responsible for oocyte activation. PLCZ1 has been identified as the factor that the sperm delivers into the egg to induce such a response. We tested the hypothesis that PLCZ1 cRNA injection can be used to activate bovine oocytes.


RESULTS
Mouse and bovine PLCZ1 cRNAs were injected into matured bovine oocytes at different concentrations. Within the concentrations tested, mouse PLCZ1 injection activated bovine oocytes at a maximum rate when the pipette concentration of cRNA ranged from 0.25 to 1 mug/muL, while bovine PLCZ1 was optimal at 0.1 mug/muL. At their most effective concentrations, PLCZ1 induced parthenogenetic development at rates similar to those observed using other activation stimuli such as Ionomycin/CHX and Ionomycin/DMAP. Injection of mouse and bovine PLCZ1 cRNA induced dose-dependent sperm-like calcium oscillations whose frequency increased over time. Injection of bovine and mouse PLCZ1 cRNA also induced IP3R-1 degradation, although bovine PLCZ1 cRNA evoked greater receptor degradation than its mouse counterpart.


CONCLUSIONS
Injection of PLCZ1 cRNA efficiently activated bovine oocytes by inducing a sperm-like calcium oscillatory pattern. Importantly, the high rate of aneuploidy encountered in parthenogenetic embryos activated by certain chemical means was not observed in PLCZ1 activated embryos.
Ross
Pablo J
PJ
Cellular Reprogramming Laboratory, Department of Animal Science, Michigan State University, East Lansing, MI 48824, USA. rosspabl@msu.edu
Beyhan
Zeki
Z
Iager
Amy E
AE
Yoon
Sook-Young
SY
Malcuit
Christopher
C
Schellander
Karl
K
Fissore
Rafael A
RA
Cibelli
Jose B
JB
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D004622:Embryo, Mammalian / Q000378:metabolism; D047108:Embryonic Development / Q000235:genetics*; D051379:Mice; D009865:Oocytes / Q000254:growth & development* / Q000378:metabolism; D010312:Parthenogenesis / Q000235:genetics*; D054801:Phosphoinositide Phospholipase C / Q000235:genetics*; D018075:RNA, Complementary; D020554:Sperm Injections, Intracytoplasmic
D018075:RNA, Complementary; D054801:Phosphoinositide Phospholipase C; C499234:Plcz1 protein, mouse; D002118:Calcium