Tight junction biogenesis in the mouse preimplantation embryo

Abstract

The mouse preimplantation embryo is a valuable model for investigating de novo formation of an epithelium. During early development, the trophectoderm differentiates and forms the wall of the blastocyst from the 32-cell stage. The tight junction (TJ) becomes fully functional at this time, resulting in blastocoel formation. Occludin, claudins and junctional adhesion molecule 1 (JAM-1) are TJ transmembrane proteins.

Here, I have analysed the expression profile of claudin-1, claudin-3 and JAM-1 by RT-PCR, immunoblotting and immunoconfocal microscopy. Claudin-1 mRNA was detected on a few occasions in late blastocysts but subsequently could not be repeated. However, claudin-1/-3 protein is first evident during cleavage at perinuclear Golgi-like sites in 16-cell embryos, followed shortly by membrane assembly at the TJ at the 32-cell stage. Antibodies that detect either claudin-1 or claudin-3 also demonstrated that these proteins may also localise to within the nucleus in earlier cleavage stages. In contrast, JAM-1 is detectable from the early 8-cell stage, earlier that any other TJ protein studied in our model. JAM-1 membrane assembly precedes E-cadherin adhesion at compaction. During compaction, JAM-1 localises predominantly to the apical microvillous pole before accumulating at cell-contact sites during later cleavage. Functional studies to investigate the role of both Claudin-1 and JAM-1 were also carried out. Claudin-1 inhibition, using a peptide corresponding to its first extracellular loop, was unsuccessful. However, neutralising JAM-1 antibody, BV11, was shown to cause a delay in embryo cavitation.

This study provides further evidence that TJ biogenesis is a multi-step process in the early embryo driven by a temporally regulated expression programme, which regulates blastocyst formation. Moreover, these data indicate that JAM-1 may have a unique role in the early stages of epithelial polarity.

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