Decidualization is a prerequisite for successful embryo implantation, in which elongated fibroblast-like endometrial stromal cells differentiate into more rounded decidual cells. Accumulating evidence has stressed the important role of the defective eutopic endometrium in infertility in endometriosis patients. However, the role of arginine methylation in the process of physiological decidualization and pathological decidualization defects is not clear. Here, we observed that the expression level of PRMT5, the main type II PRMT, was decreased in the endometrium of endometriosis patients, predominantly in stromal cells. Compared with the undecidualized state, PRMT5 was increased in the stromal cells of normal secretory endometrium in humans and in the decidua of normal pregnant mice or mice with artificially induced decidualization. The inhibition of PRMT5 resulted in a significant decrease in uterine weight and decidualization-related regulator expression, including FOXO1, HOXA10 and WNT4, in mice and IGFBP1 and prolactin levels in human endometrial stromal cells. Transcriptome analysis showed that decreased PRMT5 activity led to NF-κB signaling activation by inducing p65 translocation to the nucleus, which was also observed in endometriosis patients. Finally, overexpression of PRMT5 rescued the defective expression of IGFBP1 and prolactin in primary endometrial stromal cells from endometriosis patients. Our results indicate that promotion of PRMT5 may provide novel therapeutic strategies for the treatment of decidualization defects in infertile women, such as those with endometriosis.

Embryo adhesion is a very important step in the embryo implantation process. Homeobox A10 (HOXA10), a key transcriptional factor of endometrial receptivity, is indispensable for embryo adhesion. However, how to control the activation status of HOXA10 remains elusive. Here, we found that Mitogen-activated protein kinase kinase kinase 4 (MEKK4) was associated with HOXA10 and directly phosphorylated HOXA10 at threonine 362. This MEKK4-mediated phosphorylation enhanced HOXA10-mediated transcriptional responses and adhesion between the embryo and endometrial epithelium. Specific deletion or kinase inactivation of MEKK4 in endometrial epithelial cells attenuates adhesion between embryo and epithelium. Therefore, the identification of MEKK4 as a novel physiological positive regulator of HOXA10 activation provides mechanistic insights to improve embryo implantation success. Moreover, when Thr362 was mutated to alanine (T362A) to mimic its dephosphorylation, the protein stability and transcriptional regulation of HOXA10 were decreased. In addition, HOXA10 -promoted embryo adhesion was weakened after the mutation of Thr362, suggesting that the phosphorylation of HOXA10 at this site may be a new indicator for evaluating endometrial receptivity and judging the ‘implantation window’.

Recurrent implantation failure (RIF) is a rather thorny problem in the clinical practice of assisted reproductive technology. Due to the complex aetiology of RIF, its pathogenesis is far from fully understood, and there is no effective treatment available. Here, We explored the regulatory mechanism of the four half-domains of LIM domain 1 (FHL1), which is significantly downregulated in the endometrium of RIF patients, in blastocyst-epithelial adhesion. Indeed, FHL1 expression was dramatically increased in normal female mid-secretory endometrial epithelial cells and was abnormally reduced in RIF patients. Furthermore, FHL1 overexpression promoted blastocyst-epithelial adhesion, and interfering with FHL1 expression in the mouse uterus significantly inhibited embryo implantation. Mechanistically, FHL1 did not regulate HOXA10 mRNA expression but increased HOXA10 protein stability and activated HOXA10, thereby promoting its regulation of downstream gene expression and the β3 integrin/FAK pathway. Meanwhile, FHL1 regulates HOXA10 function by increasing HOXA10 deacetylation through enhanced binding of HOXA10 and SIRT2. SIRT2-specific inhibitors can significantly inhibit this effect. In the endometrial epithelial cells of RIF patients, the correlation between FHL1 and HOXA10 and its downstream target genes has also been verified. Finally, our data indicated FHL1 is a regulatory molecule that promotes blastocyst-epithelial adhesion. Altogether, downstream dysfunction due to aberrant FHL1 expression is an important molecular basis for embryo implantation failure in patients with RIF and to provide new potential therapeutic targets.