title: Post-fusion treatment with MG132 increases transcription factor expression in somatic cell nuclear transfer embryos in pigs.
abstract: The objective of this study was to examine the effect of post-fusion treatment of somatic cell nuclear transfer (SCNT) oocytes with the proteasomal inhibitor MG132 on maturation promoting factor (MPF) activity, nuclear remodeling, embryonic development, and gene expression of cloned pig embryos. Immediately after electrofusion, SCNT oocytes were treated with MG132 and/or caffeine for 2 hr, vanadate for 0.5 hr, or vanadate for 0.5 hr followed by MG132 for 1.5 hr. Of the MG132 concentrations tested (0-5 microM), the 1 microM concentration showed a higher rate of blastocyst formation (25.9%) than 0 (14.2%), 0.5 (16.9%), and 5 microM (16.9%). Post-fusion treatment with MG132, caffeine, and both MG132 and caffeine improved blastocyst formation (22.1%, 21.4%, and 24.4%, respectively), whereas vanadate treatment inhibited blastocyst formation (6.5%) compared to the control (11.1%). When examined 2 hr after fusion and 1 hr after activation, MPF activity remained at a higher (P < 0.05) level in SCNT oocytes that were treated post-fusion with caffeine and/or MG132, but it was decreased by vanadate. The rate of oocytes showing premature chromosome condensation was not altered by MG132 but was decreased by vanadate treatment. In addition, formation of single pronuclei was increased by MG132 compared to control and vanadate treatment. MG132-treated embryos showed increased expression of POU5F1, DPPA2, DPPA3, DPPA5, and NDP52l1 genes compared to control embryos. Our results demonstrate that post-fusion treatment of SCNT oocytes with MG132 prevents MPF degradation and increases expression of transcription factors in SCNT embryos, which are necessary for normal development of SCNT embryos.
authors.affiliation: School of Veterinary Medicine, Kangwon National University, Chunchon, Korea.
authors.forename: Jinyoung
authors.initials: J
authors.lastname: You
authors.forename: Joohyeong
authors.initials: J
authors.lastname: Lee
authors.forename: Jinyoung
authors.initials: J
authors.lastname: Kim
authors.forename: Junhong
authors.initials: J
authors.lastname: Park
authors.forename: Eunsong
authors.initials: E
authors.lastname: Lee
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000378:metabolism*; D019976:Cloning, Organism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000502:physiology; D007976:Leupeptins / Q000494:pharmacology*; D016200:Maturation-Promoting Factor / Q000378:metabolism; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors; D013552:Swine; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects* / Q000502:physiology
chemical_list: D015853:Cysteine Proteinase Inhibitors; D007976:Leupeptins; D061988:Proteasome Inhibitors; D016200:Maturation-Promoting Factor; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde