FoxL2 and Smad3 Coordinately Regulate Follistatin Gene Transcription
FoxL2 and Smad3 Coordinately Regulate Follistatin Gene Transcription
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DOI：
10.1074/jbc.m806676200
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发表时间：
2009-03-20
影响因子：
4.8
通讯作者：
Bilezikjian, Louise M.
中科院分区：
文献类型：
--
作者：
Blount, Amy L.;Schmidt, Karsten;Bilezikjian, Louise M.
卵泡抑素是一个转录靶点和激活素作用的调节剂。通过自分泌/旁分泌回路，激活素控制卵泡抑素水平，从而调节其自身的生物利用度。在促性腺激素α T3-1细胞中，激活素主要通过Smad 3在内含子Smad结合元件（SBE 1）上的作用诱导卵泡抑素转录。使用蛋白质组学方法，我们寻找参与SBE 1介导的转录的内源性α T3-1蛋白。我们确定FoxL 2，叉头家族的成员，作为SBE 1功能的候选调节剂。FoxL 2突变与以颅面缺陷和卵巢早衰为特征的小睑裂/上睑下垂/内眦赘皮综合征相关。FoxL 2定位于成年小鼠垂体的α-糖蛋白亚基和卵泡刺激素β-阳性细胞，并存在于α T3-1和L β T2细胞中，但其垂体作用在很大程度上仍然未知。我们已经确定，FoxL 2结合到位于卵泡抑素基因SBE 1位点下游的叉头结合元件（FKHB），并作为Smad 3伴侣在用激活素处理的α T3-1细胞中驱动SBE 1介导的转录。染色质免疫沉淀试验证实，内源性FoxL 2和Smad 3被募集到卵泡抑素基因的内含子增强子中，SBE 1和FKHB位点位于该基因的内含子增强子中。外源性FoxL 2增强SBE 1介导的转录，短发夹RNA介导的内源性FoxL 2蛋白的敲低在α T3-1细胞中损害这种作用。FoxL 2与Smad 3直接结合，而与Smad 2或Smad 4不结合。Smad 3和FoxL 2之间的这种关联是由Smad 3的MH 2结构域介导的，并且依赖于FoxL 2中的完整叉头结构域。总之，这些观察结果突出了FoxL 2的新作用，并表明它可能作为Smad 3靶点的转录调节因子和协调者发挥作用。
Follistatin is a transcriptional target and a modulator of activin action. Through an autocrine/paracrine loop, activin controls follistatin levels and thus regulates its own bioavailability. In gonadotropic alpha T3-1 cells, activin induces follistatin transcription primarily through the action of Smad3 at an intronic Smad-binding element (SBE1). Using a proteomics approach, we searched for endogenous alpha T3-1 proteins that participate in SBE1-mediated transcription. We identified FoxL2, a member of the forkhead family, as a candidate modulator of SBE1 function. Mutations of FoxL2 are associated with the blepharophimosis/ptosis/epicanthus inversus syndrome characterized with craniofacial defects and premature ovarian failure. FoxL2 localizes to alpha-glycoprotein subunit-and follicle-stimulating hormone beta-positive cells of the adult mouse pituitary and is present in alpha T3-1 and L beta T2 cells, but its pituitary actions remain largely unknown. We have determined that FoxL2 binds to a forkhead-binding element (FKHB) located just downstream of the SBE1 site of the follistatin gene and functions as a Smad3 partner to drive SBE1-mediated transcription in alpha T3-1 cells treated with activin. Chromatin immunoprecipitation assays confirm that endogenous FoxL2 and Smad3 are recruited to the intronic enhancer of the follistatin gene where the SBE1 and FKHB sites are located. Exogenous FoxL2 enhances SBE1-mediated transcription, and short hairpin RNA-mediated knockdown of endogenous FoxL2 protein compromises this effect in alpha T3-1 cells. FoxL2 directly associates with Smad3 but not Smad2 or Smad4. This association between Smad3 and FoxL2 is mediated by the MH2 domain of Smad3 and is dependent on an intact forkhead domain in FoxL2. Altogether, these observations highlight a novel role for FoxL2 and suggest that it may function as a transcriptional regulator and a coordinator of Smad3 targets.