Tight junction protein ZO-2 expression and relative function of ZO-1 and ZO-2 during mouse blastocyst formation

• Published in: Experimental cell research 2008-11, Vol.314 (18), p.3356-3368
• Main Authors: Sheth, Bhavwanti, Nowak, Rachael L., Anderson, Rebecca, Kwong, Wing Yee, Papenbrock, Thomas, Fleming, Tom P.
• Format: Article
• Language: English
• Subjects: 60 APPLIED LIFE SCIENCES; Animals; Blastocyst; Blastocyst - cytology; Blastocyst - drug effects; Blastocyst - metabolism; Blotting, Western; Cell Differentiation - drug effects; Cell Differentiation - genetics; Cell Differentiation - physiology; CELL MEMBRANES; CELL PROLIFERATION; Cellular biology; CLEAVAGE; Differentiation; Embryo Culture Techniques; Embryonic Development; Epithelial Cells - cytology; Epithelial Cells - physiology; EPITHELIUM; Gene expression; Gene Expression Regulation, Developmental - drug effects; Gene Expression Regulation, Developmental - physiology; GENES; INHIBITION; Membrane Proteins - antagonists & inhibitors; Membrane Proteins - genetics; Membrane Proteins - physiology; Membranes; MICE; MORPHOGENESIS; Mutation; PHENOTYPE; Phosphoproteins - antagonists & inhibitors; Phosphoproteins - genetics; Phosphoproteins - physiology; PROTEINS; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - biosynthesis; RNA, Small Interfering - pharmacology; Rodents; siRNA; Tight junction; Tight Junctions - physiology; Trophectoderm; Up-Regulation - genetics; ZO-1; ZO-2; Zonula Occludens-1 Protein; Zonula Occludens-2 Protein; ZYGOTES
• ISSN: 0014-4827; 1090-2422
• DOI: 10.1016/j.yexcr.2008.08.021

Apicolateral tight junctions (TJs) between epithelial cells are multiprotein complexes regulating membrane polarity and paracellular transport and also contribute to signalling pathways affecting cell proliferation and gene expression. ZO-2 and other ZO family members form a sub-membranous scaffold for binding TJ constituents. We investigated ZO-2 contribution to TJ biogenesis and function during trophectoderm epithelium differentiation in mouse preimplantation embryos. Our data indicate that ZO-2 is expressed from maternal and embryonic genomes with maternal ZO-2 protein associated with nuclei in zygotes and particularly early cleavage stages. Embryonic ZO-2 assembled at outer blastomere apicolateral junctional sites from the late 16-cell stage. Junctional ZO-2 first co-localised with E-cadherin in a transient complex comprising adherens junction and TJ constituents before segregating to TJs after their separation from the blastocyst stage (32-cell onwards). ZO-2 siRNA microinjection into zygotes or 2-cell embryos resulted in specific knockdown of ZO-2 mRNA and protein within blastocysts. Embryos lacking ZO-2 protein at trophectoderm TJs exhibited delayed blastocoel cavity formation but underwent normal cell proliferation and outgrowth morphogenesis. Quantitative analysis of trophectoderm TJs in ZO-2-deficient embryos revealed increased assembly of ZO-1 but not occludin, indicating ZO protein redundancy as a compensatory mechanism contributing to the mild phenotype observed. In contrast, ZO-1 knockdown, or combined ZO-1 and ZO-2 knockdown, generated a more severe inhibition of blastocoel formation indicating distinct roles for ZO proteins in blastocyst morphogenesis.