Autophagy influences maternal mRNA degradation and apoptosis in porcine parthenotes developing in vitro.
Autophagy, an essential process for cellular maintenance, cell viability, and development, is the bulk degradation of proteins and organelles. This study investigated the expression levels of autophagy-related genes and the effect of 3-methyladenine (3-MA, an autophagy inhibitor) or rapamycin (an autophagy inducer) on maternal gene degradation and apoptosis in porcine parthenotes developing in vitro. LC3, which is essential for the formation of autophagosomes, was widely expressed in porcine parthenotes. High levels of autophagy-related genes, Atg5, Beclin1 and Lc3 transcripts were expressed in the 1-cell (1C) stage and gradually decreased through the 2-cell (2C) to blastocyst stages. The mRNA expression of Gdf9, c-mos and cyclin B maintained high levels in 2C and 4-cell (4C) embryos treated with 3-MA compared with the control. The Bmp15 and cyclin B mRNA levels were significantly reduced in embryos treated with rapamycin compared with the control. These results suggest that autophagy influences the degradation of these maternal genes. Furthermore, 3-MA-treated embryos exhibited significantly reduced developmental rates, decreased total cell numbers and increased rates of apoptosis. Expression of Atg5, Beclin1 and Lc3 and synthesis of LC3 protein were significantly reduced at the blastocyst stage. Although rapamycin treatment did not affect the developmental rate, it decreased the cell number and increased the rate of apoptosis, and the expression of Atg5, Beclin1 and Lc3 and LC3 protein synthesis were increased. Finally, blastocysts derived following treatment with 3-MA or rapamycin exhibited significantly decreased expression of selected transcription factors, including Pou5f1, Sox2 and Nanog. In conclusion, our results demonstrate that autophagy influences maternal mRNA degradation and apoptosis at the blastocyst stage and suggest that autophagy plays an important role in early embryo development in the pig.
Department of Animal Sciences, Chungbuk National University, Chungbuk 361-763, South Korea.
Yong-Nan
YN
Xu
Xing-Hui
XH
Shen
Seung-Eun
SE
Lee
Jung-Suk
JS
Kwon
Deuk-Joong
DJ
Kim
Young-Tae
YT
Heo
Xiang-Shun
XS
Cui
Nam-Hyung
NH
Kim
D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D001343:Autophagy / Q000187:drug effects; D001755:Blastocyst / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D004477:Ectogenesis / Q000187:drug effects; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D059471:In Vitro Oocyte Maturation Techniques; D008540:Meiosis / Q000187:drug effects; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D009865:Oocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010312:Parthenogenesis / Q000187:drug effects; D020871:RNA Stability / Q000187:drug effects; D023121:RNA, Messenger, Stored / Q000378:metabolism*; D020123:Sirolimus / Q000494:pharmacology; D034421:Sus scrofa; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins; D008869:Microtubule-Associated Proteins; D023121:RNA, Messenger, Stored; D014157:Transcription Factors; C025946:3-methyladenine; D000225:Adenine; D020123:Sirolimus