Steroidogenic factor 1 (NR5A1) induces multiple transcriptional changes during differentiation of human gonadal-like cells

Abstract

<p>Nuclear receptor subfamily 5 group A member 1 (<em>NR5A1</em>) encodes steroidogenic factor 1 (SF1), a key regulatory factor that determines gonadal development and coordinates endocrine functions. Here, we have established a stem cell-based model of human gonadal development and applied it to evaluate the effects of <em>NR5A1</em> during the transition from bipotential gonad to testicular cells. We combined directed differentiation of human induced pluripotent stem cells (46,XY) with activation of endogenous <em>NR5A1</em> expression by conditionally-inducible CRISPR activation. The resulting male gonadal-like cells expressed several Sertoli cell transcripts, secreted anti-Müllerian hormone and responded to follicle-stimulating hormone by producing sex steroid intermediates. These characteristics were not induced without <em>NR5A1</em> activation. A total of 2691 differentially expressed genetic elements, including both coding and non-coding RNAs, were detected immediately following activation of <em>NR5A1</em> expression. Of those, we identified novel gonad-related putative <em>NR5A1</em> targets, such as SCARA5, which we validated also by immunocytochemistry. In addition, <em>NR5A1</em> activation was associated with dynamic expression of multiple gonad- and infertility-related differentially expressed genes. In conclusion, by combining targeted differentiation and endogenous activation of <em>NR5A1</em> we have for the first time, been able to examine in detail the effects of <em>NR5A1</em> in early human gonadal cells. The model and results obtained provide a useful resource for future investigations exploring the causative reasons for gonadal dysgenesis and infertility in humans.</p>