title: Tight junction protein ZO-2 expression and relative function of ZO-1 and ZO-2 during mouse blastocyst formation.
abstract: 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.
authors.affiliation: School of Biological Sciences, University of Southampton, Bassett Crescent East, Southampton SO167PX, UK.
authors.forename: Bhavwanti
authors.initials: B
authors.lastname: Sheth
authors.forename: Rachael L
authors.initials: RL
authors.lastname: Nowak
authors.forename: Rebecca
authors.initials: R
authors.lastname: Anderson
authors.forename: Wing Yee
authors.initials: WY
authors.lastname: Kwong
authors.forename: Thomas
authors.initials: T
authors.lastname: Papenbrock
authors.forename: Tom P
authors.initials: TP
authors.lastname: Fleming
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D046149:Embryo Culture Techniques; D047108:Embryonic Development; D004847:Epithelial Cells / Q000166:cytology / Q000502:physiology; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000502:physiology*; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D051379:Mice; D009154:Mutation; D010750:Phosphoproteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019108:Tight Junctions / Q000502:physiology*; D015854:Up-Regulation / Q000235:genetics; D062826:Zonula Occludens-1 Protein; D062827:Zonula Occludens-2 Protein
chemical_list: D008565:Membrane Proteins; D010750:Phosphoproteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C569043:Tjp1 protein, mouse; C569047:Tjp2 protein, mouse; D062826:Zonula Occludens-1 Protein; D062827:Zonula Occludens-2 Protein