Molecular Mechanisms Regulating Uterus Development

Project Abstract Infertility and pregnancy loss are common health disorders affecting women. Our long-term research goal is to define critical physiological and genetic pathways that regulate uterine development, function, and regeneration to diagnose,

treat, and prevent infertility and associated uterine disorders and disease. This application is specifically focused on

the glands of the uterus that are characteristic features of all uteri and critical for pregnancy. Pregnancy loss is the most common complication of human gestation, and recurrent pregnancy loss and infertility are observed in uterine gland knockout animal models. The pioneer transcription factor forkhead box A2 (FOXA2), exclusively expressed in

the glands of the mouse and human uterus, plays a pivotal role in uterine gland development and function, and is implicated in a variety of uterine pathologies (infertility, adenomyosis, endometriosis, adenocarcinoma). However, our understanding of uterine epithelial development, FOXA2 biology, and gland morphogenesis is very incomplete.

Central hypotheses of this application are that: (1) intrinsic genetic and epigenetic mechanisms govern epithelial specification, bifurcation, differentiation, and morphogenesis in the neonatal mouse uterus; (2) glandular epithelium lineage bifurcation requires Foxa2 gene activation via chromatin remodeling; and (3) FOXA2 activates gene

regulatory networks and cellular pathways critical for adenogenesis in the uterus. A team of exceptional investigators with complementary and substantial expertise in developmental biology, functional genomics, and bioinformatics will address that hypothesis by conducting a collaborative research project. Guided by strong published and preliminary

data, two specific aims are proposed: (1) understanding molecular mechanisms regulating epithelial specification and Foxa2 expression in the developing uterus; and (2) FOXA2 regulation of adenogenesis in the developing uterus. The proposed systems biology approach will integrate in vivo and in vitro studies utilizing mouse genetic models, single-

cell profiling of chromatin accessibility and gene expression (multiome), advanced bioinformatics and deep learning techniques, as well as mechanistic studies of mouse and human endometrial organoids. This approach will decipher essential conserved factors, gene regulatory networks, and cellular pathways governing epithelial development and

Foxa2 regulation and biological function. The proposed aims are conceptually and technically innovative and together will have a broad impact on the field by filling a substantial gap in our fundamental knowledge of uterine development and biology. This application specifically addresses early pregnancy loss and the genetic basis of idiopathic female

infertility, which aligns with the research priorities of the Fertility and Infertility Branch of the NICHD. Ultimately,

an increased understanding of uterine gland biology will facilitate diagnosis, prevention, and treatment of fertility and pregnancy problems, as well as benefit the development of regenerative medicine therapies aimed at organ replacement and treatments for infertility and disease in women.