Mechanisms of ovarian endocrine disruption at single-cell resolution

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Mechanisms of Ovarian Endocrine Disruption at Single-Cell Resolution Endocrine disrupting chemicals (EDCs) are exogenous chemicals that interfere with endogenous hormone synthesis, metabolism and signaling. EDC exposure during early gonadal development is suspected to play a role in ovarian dysgenesis later in life. Advancing understanding of endocrine disruption at the molecular and cellular level is essential for determining possible developmental origins of adult-onset reproductive disease. This demands sensitive in vivo models with short lifespans and innovative analytical tools to detect genetic perturbations induced by chemical exposures. The Japanese medaka fish (Oryzias latipes) is among the most studied experimental models used in EDC screening for developmental and reproductive effects. Not only is the medaka genome extensively annotated compared to other fish models, but it shares chromosomal sex determination, hormone receptor sequence homology, and sex hormone signaling axes with humans. Exposure to estrogen-contaminated water during ovarian development causes significantly delayed oocyte maturation and reduced egg production in adult medaka. However, specific mechanisms by which heterogenous cell populations of the developing ovary respond to estrogenic chemicals remain unclear. This proposal seeks to investigate how early developmental exposures to xenoestrogens alter gene regulation networks in differentiating ovarian cells, and which long-term transcriptional changes are associated with decreased female fertility. My working hypothesis is that early life stage exposures to estrogenic DDTs induce long-term transcriptional effects in ovarian somatic gonad cells, reducing fertility later in life. I will investigate this hypothesis by exposing medaka to environmentally relevant levels of o,p’- dichlorodiethyltrichloroethane (o,p’-DDT), an estrogenic isomer of the legacy organochlorine pesticide that remains a persistent public health concern. Exposures will occur during a key window of ovarian development to define organizational effects at the functional, cellular, and transcriptional levels. Aim 1 will define the long-term reproductive consequences of o,p’-DDT exposures during an early window of ovarian differentiation using functional breeding assessments and targeted quantification of estrogen- and steroidogenesis-related genes. Aim 2 will use single cell transcriptomics (scRNA-seq) to gain a comprehensive view of which ovarian somatic cell types and gene regulatory networks (both steroid hormone- dependent and hormone-independent), are permanently altered by early stage o,p’-DDT exposure. The proposed work will elucidate lasting functional and molecular changes induced by xenoestrogens during ovarian differentiation and follicular assembly in medaka, which aligns with fetal ovarian development in humans. Comprehensive identification of DDT-inducible genes may reveal novel exposure biomarkers and plausible mechanisms of endocrine and reproductive diseases.