TAF4b is a gonadal-enriched component of the general transcription complex, TFIID, and regulates a subset of granulosa-specific genes. The granulosa cells, somatic in origin, respond to a diverse set of hormonal stimuli leading to proliferation and differentiation events required for the production of a healthy oocyte. Our knock-out mouse line shows that in the absence of TAF4b, female mice develop normally with the exception of complete infertility, which stems from their inability to respond to hormonal stimuli. The present study uses an in vitro assay to test the effects of hormones on TAF4b in granulosa cells in culture, and in vivo methods to examine how loss of TAF4b affects the estrous cycle and circulating hormone levels in the mouse. TAF4b-deficient granulosa cells do not proliferate normally in response to follicle-stimulating-hormone (FSH) or estrogen, leading us to ask if the actions of these hormones act directly through TAF4b. We use the immortalized mouse granulosa cell line, NT-1, to observe any changes in TAF4b levels upon stimulation with FSH, 17-B-estradiol (E2) and the PKA activator, forskolin. NT-1 cells were grown in phenol-red-free media with 100 ng/ml FSH, 100 nM E2, 25 uM forskolin, or controls of either ethanol or PBS. Cells were harvested at multiple time-points ranging from 15 minutes to 36 hours. Western blotting of normalized protein extracts was used to assay TAF4b protein and quantitative RT-PCR was used to quantify mRNA. TAF4b shows small but statistically significant increases with all of these treatments on both the protein and mRNA levels. These data, combined with previous findings, suggest that one mechanism through which FSH and E2 act to promote normal fertility involves TAF4b directly. We previously reported that TAF4b null mice are infertile and have elevated FSH levels, and now sought to uncover the extent of disruption to their estrous cycle. Here we show that TAF4b knockout mice are in a state of persistent estrous, correlating well with the observance of high FSH levels. Vaginal smear cytology was performed every 24 hours for two weeks to assess the estrous cycle-stage of TAF4b-null mice and their littermates. Litters aged 2, 3, 4, 5 and 6 months were examined in this study. Although TAF4b-null mice are never fertile, their phenotype appears to become more severe with age. The time spent in estrous increases with age for TAF4b-null mice while their littermates continue to cycle normally. Occasionally TAF4b-null mice will exit estrous and resume a cycle. We used these instances to collect blood from pro-estrous stage mice, when E2 levels should be highest. We thought a decreased E2 concentration may accompany the elevated FSH levels seen in the knockout but, interestingly, during proestrous, knockouts have E2 serum levels comparable to wild-type age-matched mice. Together, these findings highlight TAF4b as an important partner in the hormonal regulation required for female fertility. This research was funded by NIH-T32-GM07601 and NIH-NCRR-RR015578-07.