Human CYP19A1 blocking peptide

Polycystic ovary syndrome (PCOS) is a common endocrine and metabolic disorder in reproductive women and is characterized by polycystic ovaries, hyperandrogenism and chronic anovulation. Abnormal folliculogenesis is considered as a common characteristic of PCOS. Our aim is to identify the altered circRNA expression profile in exosomes isolated from follicular fluid (FF) of PCOS patients to investigate the molecular function of exosomal circRNA, as a vital mediator in follicular microenvironment, in the aetiology and pathobiology of PCOS. In this study, the circRNA expression profile of FF exosomes were compared between PCOS and control patients by RNA sequencing (N=5 vs 5). Sixteen circRNAs showed significantly different expression. GO and KEGG pathway analyses indicated that their parental genes were enriched in PCOS-related pathways, including ovarian steroidogenesis, aldosterone synthesis and secretion, and Jak-STAT signaling. Among sixteen differentially expressed circRNAs, hsa_circ_0006877 (circLDLR) was processed from its parental LDLR (low density lipoprotein receptor) transcript, which participated in ovarian steroidogenesis. Its depletion in PCOS FF exosomes was further verified in an additional cohort (N=25 vs 25) by qRT-PCR. And a circLDLR-miR-1294-CYP19A1 competing endogenous RNA (ceRNA) network was predicted by cytoscape software, and confirmed by luciferase assay and correlative expression in the cumulus cells of PCOS patients. Mechanistically, the intercellular transfer of functional circLDLR assay and its withdrawal experiments in KGN cells showed that depleting circLDLR in exosomes increased miR-1294 expression and inhibited CYP19A1 expression in recipient cells, as well as reduced their estrogen (E2) secretion. Our findings revealed a ceRNA network of circLDLR and provided new information on abnormal follicle development in PCOS.

Follicle growth and granulosa cell health are dependent on the secretion of estradiol from granulosa cells. Estradiol is synthesized from androgen precursor by cytochrome P450 aromatase (CYP19A1), and in cattle CYP19A1 messenger RNA has a short half-life but a long (3.5kb) 3 '-untranslated region (3 ' UTR), suggesting that posttranscriptional regulation may be important for control of enzyme activity. We tested this hypothesis by inserting the CYP19A1 3 ' UTR and fragments thereof into a reporter vector between the end of the luciferase coding region and the polyadenylation signal. The full-length aromatase 3 ' UTR suppressed luciferase activity to 10% of control levels, and smaller fragments showed that this inhibitory activity lies between +926 and +1134 of the 3 ' UTR. Protein-RNA cross-linking experiments revealed that these 3 ' UTR fragments formed an RNA-protein complex of approximately 70kDa that was present in granulosa cells but not in corpus luteum, lung, liver, kidney, pancreas, or bladder extracts. The RNA-binding activity was specific to the 3 ' UTR, as shown by competition experiments with unlabeled RNA, and was present only in 3 ' UTR constructs that inhibited luciferase activity. These data suggest that posttranscriptional regulation is an important component of the control of CYP19A1 expression and involves protein binding to a specific sequence in the 3 ' UTR.