NFkB signaling and ISGylation associated with BRCA1-mutated fallopian tube epithelium

Germline BRCA1 or BRCA2 mutations (mtBRCA1 and mtBRCA2) dramatically increase risk for high-grade serous ovarian cancer (HGSOC), the most commonly diagnosed histotype. Other risk factors for this cancer, which originates primarily in the distal fallopian tube epithelium (FTE), implicate ovulation. To test whether mtBRCA1 or mtBRCA2 FTE cells respond differently to peri-ovulatory follicular fluid (FF) exposure than control patient FTE, gene expression profiles from primary FTE cultures were compared at baseline, 24h after FF exposure, and 24h after FF replacement with culture medium. Hierarchical clustering revealed both FF exposure and BRCA mutation status affect gene expression, with BRCA1 mutation having the greatest impact. Analysis revealed increased NFκB and EGFR signaling at baseline, with increased interferon signaling after recovery from FF exposure in mtBRCA1 samples. Inhibition of EGFR signaling and ISGylation by increased BRCA1 expression was verified in an immortalized FTE cell line, OE-E6/E7, stably transfected with BRCA1. Suppression of ISG15 and ISGylated protein levels by BRCA1 expression was found to be mediated by decreased NFκB signaling and was transiently suppressed by FF exposure. This study demonstrates increased NFκB signaling associated with decreased BRCA1 expression resulting in increased ISG15 and ISGylation following FF exposure, which could represent potential targets for chemoprevention. Fallopian tube epithelium from BRCA1 and BRCA2 mutation carriers and non-mutated women were seeded in collagen-IV coated transwells (n=8 for mtBRCA1, mtBRCA2 and control). The lower chamber contained cuture medium for the duration of the experiment. Upper chamber first contained culture medium, this was replaced with follicular fluid for 24h, and this was then replaced with fresh culture medium for 24h. RNA was extracted from cells after 24h FF exposure (F24), and after recovery from FF exposure (FC). Cells maintained only in culture medium were used as a control (C24).