Nuclear and nucleolar reprogramming during the first cell cycle in bovine nuclear transfer embryos.
The immediate events of genomic reprogramming at somatic cell nuclear transfer (SCNT) are to high degree unknown. This study was designed to evaluate the nuclear and nucleolar changes during the first cell cycle. Bovine SCNT embryos were produced from starved bovine fibroblasts and fixed at 0.5, 1, 2, 3, 4, 8, 12, and 16 h postactivation (hpa). Parthenogenetic (PA) embryos were used as control. The SCNT and PA embryos were processed for lacmoid staining, autoradiography, transmission electron microscopy (TEM), and immunofluorescence localization of: upstream binding factor (UBF) and fibrillarin at 4 and 12 hpa. Likewise, starved and nonstarved fibroblasts were processed for autoradiography and TEM. The fibroblasts displayed strong transcriptional activity and active fibrillogranular nucleoli. None of the reconstructed embryos, however, displayed transcriptional activity. In conclusion, somatic cell nuclei introduced into enucleated oocytes displayed chromatin condensation, partial nuclear envelope breakdown, nucleolar desegregation and transcriptional quiescence already at 0.5 hpa. Somatic cell cytoplasm remained temporally attached to introduced nucleus and nucleolus was partially restored indicating somatic influence in the early SCNT phases. At 1-3 hpa, chromatin gradually decondensed toward the nucleus periphery and nuclear envelope reformed. From 4 hpa, the somatic cell nucleus gained a PN-like appearance and displayed NPBs suggesting ooplasmic control of development.
Østrup
Olga
O
Constantin the Philosopher University, Nitra, Slovak Republic. osvarcova@gmail.com
Petrovicova
Ida
I
Strejcek
Frantisek
F
Morovic
Martin
M
Lucas-Hahn
Andrea
A
Lemme
Erika
E
Petersen
Bjorn
B
Niemann
Heiner
H
Laurincik
Jozef
J
Maddox-Hyttel
Poul
P
D000818:Animals; D002417:Cattle; D002453:Cell Cycle / Q000502:physiology*; D002466:Cell Nucleolus / Q000378:metabolism*; D002478:Cells, Cultured; D042002:Chromatin Assembly and Disassembly / Q000502:physiology*; D002868:Chromosomal Proteins, Non-Histone / Q000096:biosynthesis; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental / Q000502:physiology; D009685:Nuclear Envelope / Q000378:metabolism; D053652:Nuclear Transfer Techniques; D035463:Pol1 Transcription Initiation Complex Proteins / Q000096:biosynthesis
D002868:Chromosomal Proteins, Non-Histone; D035463:Pol1 Transcription Initiation Complex Proteins; C047458:fibrillarin; C063844:transcription factor UBF