Elucidating the Role of Hox Transcription Factors in the Chromatin Context

许多转录因子（tf）在多种细胞类型中表达，但它们在不同的细胞环境中以最高的精度控制不同的转录程序。这种广泛表达但具有特异性作用的tf最突出的例子之一是Hox蛋白。有趣的是，尽管Hox tf在许多细胞类型中都很活跃，但它可以“重编程”细胞的区域身份，这是在动物的前后轴上受到强烈的表观遗传控制的。这种行为表明Hox蛋白可以诱导染色质打开，使它们能够适应谱系特异性基因表达程序以适应区域要求。然而，Hox蛋白在体内调控局部染色质状态和结构中的作用及其潜在机制仍然知之甚少。基于我们最近的发现，Hox TF Ubx以细胞谱系限制的方式与Brahma染色质重塑复合体的组分相互作用，以及Hox TF结合开放和关闭染色质在区域和谱系命运调节中发挥双重作用的事实，我们现在将探索Hox TF先驱染色质的可能性。我们假设Hox染色质先锋依赖于与核小体上低亲和力DNA基序的结合，并且通过与谱系限制性（先锋）tf相互作用，将一般细胞谱系程序调整为区域特异性需求。为了严格检验我们的假设，我们将解决Hox结合的模式，序列要求和与其他（先驱）tf的功能相互作用。为此，我们将采用最先进的基因组技术，这将使我们能够同时探测TF染色质结合和体内局部染色质环境，而不需要结合位点预测，以及补充生化和遗传方法。我们将使用Ubx及其谱系特异性相互作用伙伴Tin和Grh以及通用相互作用伙伴Exd作为模型，它们已被证明可以增加Hox与封闭染色质的结合，并将使用体外、细胞和体内方法研究它们在染色质打开中的作用。总之，这项工作将有助于解决发育生物学中的一个主要难题，即Hox tf如何在分配区域身份的同时高度特异性地发挥作用，同时实现细胞系稳定的一般功能。

Many transcription factors (TFs) are expressed in multiple cell types, yet they control distinct transcriptional programs with highest precision in different cellular contexts. One of the most prominent examples of such broadly expressed but specifically acting TFs are the Hox proteins. Intriguingly, Hox TFs, despite being active in many cell types, can “reprogram” the regional identity of cells, which is under strong epigenetic control along the anterior-posterior axis of animals. This behavior suggests that Hox proteins can induce chromatin opening, allowing them to adapt lineage-specific gene expression programs to the regional requirements. However, the role of Hox proteins in the regulation of local chromatin state and configuration in vivo and the underlying mechanisms are still poorly understood. Building on our recent findings that the Hox TF Ubx interacts in a cell lineage restricted fashion with components of the Brahma chromatin remodelling complex and the fact that Hox TFs bind to open and closed chromatin to carry out dual roles in regional and lineage fate regulation, we will now explore the possibility that Hox TFs pioneer chromatin. We hypothesize that Hox chromatin pioneering is dependent on binding to low-affinity DNA motifs on nucleosomes and that generic cell lineage programs are tuned to region-specific requirements by interacting with lineage-restricted (pioneer) TFs. In order to rigorously test our hypothesis, we will resolve the modalities of Hox binding, the sequence requirements and the functional interaction with other (pioneer) TFs. To this end, we will employ state of the art genomic techniques, which will allow us to simultaneously probe TF chromatin binding and the local chromatin environment in vivo, without the need of binding site predictions, as well as complementing biochemical and genetic approaches. We will use Ubx and its lineage-specific interaction partners Tin and Grh as well as the generic interaction partner Exd, which has been shown to increase Hox binding to closed chromatin, as models, and will study their role in chromatin opening using in vitro, in cellulo and in vivo approaches. In sum, this work will be instrumental in resolving one of the major puzzles in Developmental Biology, which is how Hox TFs can function highly specifically in assigning regional identity while fulfilling at the same time generic functions in cell lineage stabilization.