Mitochondrial physiology and gene expression analyses reveal metabolic and translational dysregulation in oocyte-induced somatic nuclear reprogramming.
While reprogramming a foreign nucleus after somatic cell nuclear transfer (SCNT), the enucleated oocyte (ooplasm) must signal that biomass and cellular requirements changed compared to the nucleus donor cell. Using cells expressing nuclear-encoded but mitochondria-targeted EGFP, a strategy was developed to directly distinguish maternal and embryonic products, testing ooplasm demands on transcriptional and post-transcriptional activity during reprogramming. Specifically, we compared transcript and protein levels for EGFP and other products in pre-implantation SCNT embryos, side-by-side to fertilized controls (embryos produced from the same oocyte pool, by intracytoplasmic injection of sperm containing the EGFP transgene). We observed that while EGFP transcript abundance is not different, protein levels are significantly lower in SCNT compared to fertilized blastocysts. This was not observed for Gapdh and Actb, whose protein reflected mRNA. This transcript-protein relationship indicates that the somatic nucleus can keep up with ooplasm transcript demands, whilst transcription and translation mismatch occurs after SCNT for certain mRNAs. We further detected metabolic disturbances after SCNT, suggesting a place among forces regulating post-transcriptional changes during reprogramming. Our observations ascribe oocyte-induced reprogramming with previously unsuspected regulatory dimensions, in that presence of functional proteins may no longer be inferred from mRNA, but rather depend on post-transcriptional regulation possibly modulated through metabolism.
Max-Planck Institute for Molecular Biomedicine, Münster, Germany.
Telma C
TC
Esteves
Olympia E
OE
Psathaki
Martin J
MJ
Pfeiffer
Sebastian T
ST
Balbach
Dagmar
D
Zeuschner
Hiroshi
H
Shitara
Hiromichi
H
Yonekawa
Marcin
M
Siatkowski
Georg
G
Fuellen
Michele
M
Boiani
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000235:genetics / Q000502:physiology*; D020285:Cryoelectron Microscopy; D005260:Female; D006861:Hydrogen Peroxide / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000235:genetics / Q000276:immunology / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D046529:Microscopy, Electron, Transmission; D008928:Mitochondria / Q000378:metabolism* / Q000648:ultrastructure; D009865:Oocytes / Q000166:cytology* / Q000648:ultrastructure
D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D002118:Calcium