PROJECT ABSTRACT Infertility affects up to 13% of childless couples yet the biological mechanisms underlying infertility and the effects of infertility on overall health remains poorly understood. The goal of Project 1 within The Massachusetts General Hospital Harvard Center for Reproductive Medicine will be to apply human genomics, transcriptomics and phenomics to understand the mechanistic biological drivers of rare disorders of human infertility. A pervasive methodologic theme of 'omics’ technologies runs through Project 1’s three specific aims. In Aim 1, infertility will be viewed through the prism of rare hypogonadotropic and hypergonadotropic conditions that cause infertility. Clinical investigators will apply contemporary genomic techniques to define the underlying genetic architecture of infertility and to identify master regulatory pathways and networks that determine fertility. Three unique clinical cohorts will be utilized to achieve this aim: US-based admixed cohorts of patients with idiopathic hypogonadotropic hypogonadism (IHH)/Kallmann Syndrome [Massachusetts General Hospital] and primary ovarian insufficiency [University of Utah]; and, a Saudi Arabia-based consanguineous cohort of patients with a spectrum of rare Mendelian forms of infertility. The full spectrum of genetic variation (coding and non-coding single nucleotide variants, insertion/deletion variants and structural variants) that confer substantial relative risk for infertility will be determined and causal genes identified will be coalesced into common, final pathways elucidating the predominant drivers of rare forms of infertility. In Aim 2, genetic variants identified from Aim 1 and variants identified in Project 2 of the Center that relate to hypothalamic-hypogonadotropic forms of infertility will be validated in CRISPR-engineered GnRH neurons derived from induced pluripotent stem cells generated by the Genomics & Functional Core of the Center. Specifically, the cellular and molecular consequences of genetic variation on GnRH neurons will be defined by comparing and contrasting the GnRH transcriptome, morphology, migratory capability and secretory function between wild-type and edited GnRH neurons. Similarly, genetic variants relating to hypergonadotropic forms of infertility leading to primary ovarian insufficiency will be studied in a Drosophila model system by ovary-specific RNAi knockdown (or overexpression) experiments. Finally, in Aim 3, a hospital-based human biobank (Partners Biobank) will be utilized to perform a recall-by-genotype based targeted phenotypic evaluation in individuals harboring pathogenic variants in infertility-associated genes. The full reproductive phenotype (“reproductome”) will be defined using deep phenotyping studies that will define the effects of harboring genetic risk variants on GnRH-induced pituitary LH pulse profiles and hypothalamic-pituitary responsiveness to exogenous kisspeptin administration. Through these coordinated studies, this project will aid in the diagnosis and management of infertility, inform genetic risk prediction for infertility conditions, and facilitate the development of novel therapeutic options for infertility.