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Expression of ZO-1 and occludin at mRNA and protein level during preimplantation development of the pig parthenogenetic diploids

Published online by Cambridge University Press:  22 February 2011

Shangdan Xu ,
Jibak Lee  and
Masashi Miyake
Shangdan Xu
Affiliation:
Department of Bioresource Sciences, Graduate School of Science and Technology, Kobe University, Nada-ku, Kobe 657–8501, Japan.
Jibak Lee
Affiliation:
Chromosome Dynamics Laboratory, RIKEN Advanced Science Institute, Wako, Saitama 351–0198, Japan.
Masashi Miyake*
Affiliation:
Department of Bioresource Sciences, Graduate School of Science and Technology, Kobe University, Nada-ku, Kobe 657–8501, Japan. Organization of Advanced Science and Technology, Kobe University, Nada-ku, Kobe 657–8501, Japan.
*
All correspondence to: Masashi Miyake. Department of Bioresource Sciences, Graduate School of Science and Technology, Kobe University, Nada-ku, Kobe 657–8501, Japan. Tel: +81–78–803–6581. Fax: +81–78–803–5807. e-mail address: [email protected]
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Summary

Expression of mRNAs and proteins of ZO-1 and occludin was analyzed in pig oocytes and parthenogenetic diploid embryos during preimplantation development using real-time RT-PCR, western blotting and immunocytochemistry. All germinal vesicle (GV) and metaphase (M)II oocytes and preimplantation embryos expressed mRNAs and proteins of ZO-1 and occludin. mRNA levels of both ZO-1 and occludin decreased significantly from GV to MII, but increased at the 2-cell stage followed by temporal decrease during the early and late 4-cell stages. Then, both mRNAs increased after compaction. Relative concentration of zo1α− was highest in 2-cell embryos, while zo1α+ was expressed from the morula stage. Occludin expression greatly increased after the morula stage and was highest in expanded blastocysts. Western blotting analysis showed constant expression of ZO-1α− throughout preimplantation development and limited translation of ZO-1α+ from the blastocysts, and species-specific expression pattern of occludin. Immunocytochemistry analysis revealed homogeneous distribution of ZO-1 and occludin in the cytoplasm with moderately strong fluorescence in the vicinity of the contact region between blastomeres, around the nuclei in the 2-cell to late 4-cell embryos, and clear network localization along the cell-boundary region in embryos after the morula stage. Present results show that major TJ proteins, ZO-1 and occludin are expressed in oocytes and preimplantation embryos, and that ZO-1α+ is transcribed by zygotic gene activation and translated from early blastocysts with prominent increase of occludin at the blastocyst stage.

Keywords

Parthenogenetic embryoPigPreimplantationTight junction
Type
Research Article
Information
Zygote , Volume 20 , Issue 2 , May 2012 , pp. 147 - 158
Copyright
Copyright © Cambridge University Press 2011

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