Among nearly 100 mammalian species, implantation can be suspended at blastocyst stage for a certain time and reactivated under favorable conditions, a phenomenon known as embryonic diapause. Until now, the underlying molecular mechanism governing embryonic diapause and reactivation for implantation remained largely unknown. Here we conducted the first integral proteomic analysis of blastocysts from diapause to reactivation by using a physiologically relevant mouse delayed implantation model. More than 6000 dormant and reactivated blastocysts were used for the proteomic analysis. A total of 2255 proteins were detected. Various cellular and molecular processes, including protein translation, aerobic glycolysis, pentose phosphate pathway, purine nucleotide biosynthesis, glutathione metabolism, and chromatin organization were identified as differentially regulated. In particular, we demonstrated a remarkable activation of mitochondria in blastocysts upon reactivation from dormancy, highlighting their essential physiological significance. Moreover, the activities of the endosome-lysosome system were prominently enhanced in the mural trophectoderm of reactivated blastocysts, accompanied by active phagocytosis at the fetal-maternal interface, suggesting a critical role in promoting trophoblast invasion. Collectively, we provided an integral proteomic view upon the regulatory network of blastocyst reactivation from diapause, which will help to better interpret the nature of embryonic diapause and reactivation in wild animals and to identify molecular indicators for selecting blastocysts with high implantation competency.

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Estrogen and progesterone coupled with locally produced signaling molecules are essential for embryo implantation. However, the hierarchical landscape of the molecular pathways that governs this process remains largely unexplored. Here we show that the protein tyrosine phosphatase Shp2, a positive transducer of RTK signaling, is predominately localized in the nuclei in the periimplantation mouse uterus. Uterine-specific deletion of Shp2 exhibits reduced progesterone receptor (PR) expression and progesterone resistance, which derails normal uterine receptivity, leading to complete implantation failure in mice. Notably, the PR expression defects are attributed to the limited estrogen receptor α (ERα) activation in uterine stroma. Further analysis reveals that nuclear Shp2, rather than cytosolic Shp2, promotes the ERα transcription activity. This function is achieved by enhancing the Src kinasemediated ERα tyrosine phosphorylation, which facilitates ERα binding to Pgr promoter in an ERK-independent manner in periimplantation uteri. Besides uncovering a regulatory mechanism, this study could be clinically relevant to dysfunctional ERα-caused endometrial disorders in women.Shp2 | ERK signaling | Src kinase | estrogen receptor | uterine receptivity S uccessful implantation requires synchronization between an implantation-competent blastocyst and a receptive uterus. In humans, natural conception per cycle is poor (∼30%), and ∼75% of failed pregnancies are considered to be due to implantation failure (1). One-third of implantation failure is attributed to the embryo itself, whereas the remaining two-thirds appear to result from inadequate uterine receptivity (2). The endometrium enters into a receptive stage for blastocyst implantation only in a restricted time period termed "implantation window," which is dominated by precisely regulated proliferation and differentiation of endometrial epithelium and stroma under the influence of progesterone and estrogen (3).Estrogen and progesterone bind to the estrogen receptor (ER) and progesterone receptor (PR), respectively, coupled with specific cofactors to endure the optimal functions. The activity of these nuclear receptors is also regulated at the posttranslational level by various modifications, such as phosphorylation, which can influence the protein stability, interaction with the cofactors and DNA binding affinity. So far, a wide range of nuclear receptor cofactors have been identified to ensure the normal ER transcriptional activation, such as the well-known steroid receptor coactivator (SRC) family members, which bind with ERα in the chromatin to recruit the histone acetyltransferase p300 (P300) for transcriptional activation (4-6). It is conceivable that ordered and combinatorial recruitment of cofactors at a specific target promoter after ERα binding to DNA sequence was essential to ensure target gene transcription. Recent evidence shows that nuclear receptor coactivator 6 is essential for embryo implantation by maintaining the appropriate level and activity of uteri...

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