Effect of trychostatin A treatment on gene expression in cloned mouse embryos.
Histone deacetylation occurs upon the transfer of somatic nuclei into enucleated oocytes, but its role in reprogramming somatic chromatin to the totipotent state is unknown. To investigate the importance of histone deacetylation in reprogramming, we constructed embryos by electrofusing breast cancer cells with enucleated mouse oocytes. The reconstructed embryos were then cultured before and/or after activation for 6h in the presence of trychostatin A (TSA), a potent inhibitor of histone deacetylase. Total RNA was isolated from these TSA-treated and untreated embryos and real-time reverse transcription PCR was conducted to monitor transcription of ErbB2, Muc1, eIF-4C, MuERV-L, and c-mos genes. The nuclear-cytoplasmic interaction inhibited typical expression of ErbB2 and Muc1 in the somatic cells. Moreover, the inhibition of histone deacetylation prior to activation did not increase the levels of eIF-4C, MuERV-L, and c-mos expression in the nuclear transfer (NT) embryos (P>0.05), whereas additional treatment with 100nM TSA beyond the activation point improved expression of these genes (P<0.05). Trychostatin A treatment also improved the development rates of NT embryos at the 2-cell, 4-cell, and blastocyst stages (78.6% vs. 90.2%, 45.2% vs. 68.9%, and 16.7% vs. 30.3%, respectively, P<0.05). We hypothesized that the reprogramming of gene expression in NT embryos is independent of somatic histone deacetylation, and that hyperacetylation may have a positive effect on NT embryo development.
Shao
G-B
GB
College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095, People's Republic of China.
Ding
H-M
HM
Gao
W-L
WL
Li
S-H
SH
Wu
C-F
CF
Xu
Y-X
YX
Liu
H-L
HL
D000818:Animals; D001943:Breast Neoplasms / Q000473:pathology; D002467:Cell Nucleus / Q000473:pathology; D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000187:drug effects; D019976:Cloning, Organism / Q000379:methods; D046149:Embryo Culture Techniques; D004622:Embryo, Mammalian / Q000187:drug effects* / Q000378:metabolism; D047108:Embryonic Development / Q000187:drug effects / Q000235:genetics; D005260:Female; D005307:Fertilization in Vitro; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D006877:Hydroxamic Acids / Q000494:pharmacology*; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D053652:Nuclear Transfer Techniques; D011247:Pregnancy
D006877:Hydroxamic Acids; C012589:trichostatin A