Bone Morphogenetic Protein 2 Acts via Inhibitor of DNA Binding Proteins to Synergistically Regulate Follicle-Stimulating Hormone β Transcription with Activin A

Abstract

We recently reported that bone morphogenetic proteins (BMPs) 2 and 4 can stimulate FSHbeta-subunit (Fshb) transcription alone and in synergy with activins. We further showed that BMP2 signals via the BMP type IA receptor (or activin receptor-like kinase 3) to mediate its effects. However, the intracellular mechanisms through which BMP2 regulates Fshb are unknown. In the current study, we used cDNA microarray analyses (and validation by real-time quantitative RT-PCR) to identify BMP2 target genes in the murine gonadotrope cell line, LbetaT2. Short-interfering RNA-mediated knockdown, overexpression, and coimmunoprecipitation experiments were used to examine the potential functional roles of selected gene products. Quantitative RT-PCR analysis largely confirmed the results of the array analyses, and inhibitors of DNA binding 1, 2, and 3 (Id1, Id2, and Id3) were selected for functional analyses. Knockdown of endogenous Id2 or Id3, but not Id1, diminished the synergistic effects of BMP2 and activin A on Fshb transcription. Overexpression of Id1, Id2, or Id3 alone had no effect, but all three potentiated activin A or mothers against decapentaplegic homolog (SMAD)3 induction of Fshb transcription. Though the precise mechanism through which Ids produce their effects are not yet known, we observed physical interactions between Id1, Id2, or Id3 and SMAD3. Collectively, the data suggest that BMP2 synergistically regulates Fshb transcription with activins, at least in part, through the combined actions of Ids 2 or 3 and SMAD3.