How homeodomain proteins gain sufficient specificity to control different cell fates has been a long-standing problem in developmental biology. The conserved Gsx homeodomain proteins regulate specific aspects of neural development in animals from flies to mammals, and yet they belong to a large transcription factor family that bind nearly identical DNA sequences in vitro. Here, we show that the mouse and fly Gsx factors unexpectedly gain DNA binding specificity by forming cooperative homodimers on precisely spaced and oriented DNA sites. High-resolution genomic binding assays revealed that Gsx2 binds both monomer and homodimer sites in the developing mouse ventral telencephalon. Importantly, reporter assays showed that Gsx2 mediates opposing outcomes in a DNA binding site-dependent manner: Monomer Gsx2 binding represses transcription, whereas homodimer binding stimulates gene expression. In Drosophila, the Gsx homolog, Ind, similarly represses or stimulates transcription in a site-dependent manner via an autoregulatory enhancer containing a combination of monomer and homodimer sites. Integrating these findings, we test a model showing how the homodimer to monomer site ratio and the Gsx protein levels defines gene up-regulation versus down-regulation. Altogether, these data serve as a new paradigm for how cooperative homeodomain transcription factor binding can increase target specificity and alter regulatory outcomes.

中文翻译：

---

保守的 Gsx2/Ind 同源域单体与同源二聚体 DNA 结合定义了果蝇和小鼠的调节结果

同源域蛋白如何获得足够的特异性来控制不同的细胞命运一直是发育生物学中的一个长期问题。保守的 Gsx 同源域蛋白调节从果蝇到哺乳动物的动物神经发育的特定方面，但它们属于一个大型转录因子家族，在体外结合几乎相同的 DNA 序列。在这里，我们展示了鼠标和果蝇 Gsx 因子通过在精确间隔和定向的 DNA 位点上形成合作同源二聚体，意外地获得了 DNA 结合特异性。高分辨率基因组结合测定显示 Gsx2 结合发育中的小鼠腹侧端脑中的单体和同源二聚体位点。重要的是，报告基因检测表明 Gsx2 以 DNA 结合位点依赖性方式介导相反的结果：单体 Gsx2 结合抑制转录，而同源二聚体结合刺激基因表达。在果蝇，Gsx 同源物，Ind，类似地通过包含单体和同源二聚体位点组合的自动调节增强子以位点依赖性方式抑制或刺激转录。整合这些发现，我们测试了一个模型，显示同二聚体与单体位点的比率和 Gsx 蛋白水平如何定义基因上调与下调。总而言之，这些数据为协同同源域转录因子结合如何提高目标特异性和改变调节结果提供了新的范例。