Preimplantation development of mouse oocytes activated by different levels of human phospholipase C zeta.
BACKGROUND
A sperm-specific phospholipase C zeta (PLCzeta) has been shown to trigger Ca(2+) oscillations in mouse and human oocytes and appears to be the sperm factor responsible for activation at fertilization. Previously, complementary RNA (cRNA) injection was used to introduce PLCzeta into oocytes, but it was unclear how much PLCzeta protein is required for development. Here we have injected cRNA encoding luciferase-tagged human PLCzeta (hPLCzeta-luc) into mouse oocytes and established the relationship between hPLCzeta-luc expression, Ca(2+) oscillations and development.


METHODS
Mouse oocytes were injected with hPLCzeta-luc cRNA and a fluorescent Ca(2+)dye to monitor hPLCzeta-luc expression and Ca(2+) oscillations, respectively. After inducing diploidy, development in vitro was monitored in hPLCzeta-luc cRNA microinjected oocytes and compared with parallel oocytes activated by incubation in Sr(2+).


RESULTS
Repetitive Ca(2+) oscillations and oocyte activation were triggered by hPLCzeta over a wide range of luciferase expression levels. However, subsequent development of embryos to the blastocyst stage was observed only when expression of hPLCzeta-luc was optimized within a specific range. The blastocyst cell number was also affected by the level of hPLCzeta expression.


CONCLUSIONS
Human PLCzeta can readily activate mouse oocytes, however, effective development to blastocyst stages is only achieved within a specific window of hPLCzeta-luc protein expression levels.
Yu
Y
Y
Department of Obstetrics and Gynaecology, School of Medicine, Cardiff University, Heath Park, Cardiff CF14 4XN, UK.
Saunders
C M
CM
Lai
F A
FA
Swann
K
K
D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D002452:Cell Count; D005260:Female; D018014:Gene Transfer Techniques; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009865:Oocytes / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D009991:Oscillometry; D054801:Phosphoinositide Phospholipase C / Q000235:genetics / Q000494:pharmacology*
D008156:Luciferases; C499235:PLCZ1 protein, human; D054801:Phosphoinositide Phospholipase C; D002118:Calcium