Loss of HOXA10 activates NLRP3 for epithelial plasticity and pyroptosis in endometrium during embryo implantation.

Embryo implantation requires transient modulation of epithelial integrity at the embryo- endometrium interface, yet the molecular mechanisms that govern epithelial remodelling remain incompletely understood. In our previous work, we demonstrated that the luminal epithelial cells undergo a partial epithelial-to-mesenchymal transition (pEMT) at the site of embryo implantation, regulated by the homeobox transcription factor HomeoboxA10 (HOXA10). Here, we identify NOD-like receptor family pyrin domain-containing 3 (NLRP3), a key inflammasome component, as a direct downstream effector of HOXA10 that is essential for luminal epithelial cell remodelling. CUT&RUN profiling revealed that HOXA10 binds to regulatory regions of several NLRP family members, including NLRP3, and loss of HOXA10 in vivo results in increased NLRP3 expression in the luminal epithelial cells. Single-cell RNA-seq and immunostaining confirmed that NLRP3 is specifically upregulated in luminal epithelial cells undergoing pEMT at the time of implantation. This was accompanied by co-localization of NLRP3 with apoptosis-associated speck-like protein containing a CARD (caspase activation and recruitment domain) and CASP1 in luminal epithelial cells, suggesting inflammasome activation at the time of implantation. Crucially, treatment with NLRP3 inhibitor (MCC950) on the day of implantation, impeded pEMT and blocked pyroptosis mediated by gasdermin D in the luminal epithelial cells, leading to the retention of luminal epithelium at the site of implantation and ultimately impairing embryo invasion. Our findings collectively show that HOXA10 preserves epithelial identity by inhibiting NLRP3, and its downregulation promotes NLRP3 mediated inflammasome production and pyroptosis, allowing for epithelial clearing for a successful implantation. This study demonstrates the dual function of NLRP3 in inducing epithelial plasticity and cell death at the embryo implantation site, highlighting a precisely regulated inflammatory mechanism.