Tight junction protein ZO-2 expression and relative function of ZO-1 and ZO-2 during mouse blastocyst formation.
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.
Sheth
Bhavwanti
B
School of Biological Sciences, University of Southampton, Bassett Crescent East, Southampton SO167PX, UK.
Nowak
Rachael L
RL
Anderson
Rebecca
R
Kwong
Wing Yee
WY
Papenbrock
Thomas
T
Fleming
Tom P
TP
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
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