ABSTRACT: In order for the human uterus to become receptive to an implanting embryo, the endometrial stromal cells must undergo a complex proliferation and differentiation response to ovarian hormones, termed decidualization. Despite the integral role of uterine receptivity in the implantation phase of pregnancy, and despite many expensive in vitro fertilization procedures proving unsuccessful due to uterine deficiency, the molecular pathways underlying decidualization remain woefully understudied. One major development in the study of decidualization was the generation of immortalized human endometrial stromal cells (hESCs), which respond transcriptionally robustly to the ovarian hormones progesterone and estrogen. As preliminary data to the proposed study, we generated a reporter cell line sub-cloned from hESCs that expresses YFP under the control of the prolactin promoter, and therefore exhibits a consistent and robust induction of YFP when treated with a cocktail of ovarian hormones. These cells, termed hESC-PRLYs, represent a powerful screening tool to assess decidualization under various genomic and environmental stresses. Using these cells we performed a full genome siRNA screen to map the human decidualization genetic network, and found that among the most influential gene families required for normal decidualization are homeodomain transcription factors. The proposed study aims to first map the temporal induction profile of homeodomain proteins in hESCs in response to hormone using qPCR. Next, the requirement of homeodomain proteins in decidualization will be determined by generating conditional knockout mice using Pgr-cre for the top candidate factors. Knockout mice will undergo assessment including fertility study with endpoint of pups per litter, gestational monitoring, blastocyst flush, implantation assay, and artificial decidualization assay wherever appropriate to determine whether female fertility is affected by the loss of each gene, and at which point in the process fertility fails. The candiate gene determined to have the most impact on decidualization will undergo downstream assessment to map its targets in the decidua. This will include qPCR of potential targets in murine knockout uterus, and if time permits, RNA sequencing of mutant murine pregnant uterine stroma, Crispr knockout (or siRNA knockdown if Crispr fails) hESCs, and SpDAM assay in hESC unbiasedly identify downstream affected genes and overlap human and murine datasets. Together these data will define the robust role of homeodomain transcription factors in decidualization, and identify one or more major targets for potential pharmacological modulation in the treatment of female implantation-level infertility. Access to the newest technologies, insightful lectures, regular symposia and the fostering of a collaborative atmosphere will be an integral part of the training process. Washington University provides an unparalleled environment of advisement by the foremost ...