BACKGROUND FoxH1 is a forkhead transcription factor with conserved key functions in vertebrate mesoderm induction and left-right patterning downstream of the TGF-beta/Nodal signaling pathway. Binding of the forkhead domain (FHD) of FoxH1 to a highly conserved proximal sequence motif was shown to regulate target gene expression. RESULTS We identify the conserved microRNA-430 family (miR-430) as a novel target of FoxH1. miR-430 levels are increased in foxH1 mutants, resulting in a reduced expression of transcripts that are targeted by miR-430 for degradation. To determine the underlying mechanism of miR-430 repression, we performed chromatin immunoprecipitation studies and overexpression experiments with mutant as well as constitutive active and repressive forms of FoxH1. Our studies reveal a molecular interaction of FoxH1 with miR-430 loci independent of the FHD. Furthermore, we show that previously described mutant forms of FoxH1 that disrupt DNA binding or that lack the C-terminal Smad Interaction Domain (SID) dominantly interfere with miR-430 repression, but not with the regulation of previously described FoxH1 targets. CONCLUSIONS We were able to identify the distinct roles of protein domains of FoxH1 in the regulation process of miR-430. We provide evidence that the indirect repression of miR-430 loci depends on the connection to a distal repressive chromosome environment via a non-canonical mode. The widespread distribution of such non-canonical binding sites of FoxH1, found not only in our study, argues against a function restricted to regulating miR-430 and for a more global role of FoxH1 in chromatin folding.

中文翻译：

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FoxH1通过非规范性调控抑制斑马鱼早期胚胎发育中的miR-430。

背景FoxH1是叉头转录因子，在脊椎动物中胚层诱导和TGF-beta / Nodal信号通路下游的左右模式中具有保守的关键功能。FoxH1的叉头域（FHD）与高度保守的近端序列基序的结合显示出可调节靶基因的表达。结果我们确定保守的microRNA-430家族（miR-430）是FoxH1的新型靶标。在foxH1突变体中，miR-430水平升高，导致miR-430靶向降解的转录本表达降低。为了确定miR-430抑制的潜在机制，我们进行了染色质免疫沉淀研究和过表达实验，研究对象为FoxH1的突变型以及组成型活性和抑制型。我们的研究揭示了FoxH1与独立于FHD的miR-430基因座的分子相互作用。此外，我们表明破坏DNA结合或缺乏C端Smad相互作用域（SID）的先前描述的FoxH1突变形式主要干扰miR-430抑制，但不影响先前描述的FoxH1靶标的调控。结论我们能够确定FoxH1蛋白结构域在miR-430调控过程中的独特作用。我们提供的证据表明，miR-430基因座的间接抑制取决于通过非规范模式与远端抑制性染色体环境的连接。FoxH1这类非规范结合位点的广泛分布，不仅在我们的研究中发现，