Identification and characterization of FGF target genes

为了解决我们对哪些基因响应FGF信号传导的知识不足，我们正在进行一个项目，从分子上定义FGF靶基因，以及它们的功能和调节。在过去的工作中，我们和其他人已经确定了由Gbx 1和Gbx 2编码的转录因子作为FGF靶点。最近，我们研究了Gbx家族成员之间的功能关系，在发展中的脊髓使用组合Gbx小鼠突变体。我们发现，如果另一个不存在，每个Gbx基因都会上调。此外，Gbx基因调节PAX 2+背侧抑制性中间神经元亚群的发育。此外，在Gbx突变体中，增殖细胞扩展到解剖学定义的套区。最后，我们的数据显示，在胚胎中期，Gbx突变体的腹侧脊髓中的凋亡细胞死亡显著增加。虽然我们的研究表明，Gbx基因家族的两个成员都参与PAX 2+背侧中间神经元亚群的发育和腹侧运动神经元的存活，但Gbx 1和Gbx 2不足以在遗传上弥补彼此的损失。因此，我们的研究为Gbx 1和Gbx 2在脊髓发育中的关系提供了新的见解（J Dev Biol.2020）。PMID：32244588）。在目前的工作中，我们证明了Hes 7转录抑制因子显然是FGF 4信号转导的直接靶点。在脊椎动物发育过程中，体节前中胚层（PSM）周期性地分节，体节将形成分节的脊柱和相关的肌肉、结缔组织和真皮。体节发生的周期性受振荡Notch活性的节段时钟调节。我们研究了仅缺乏Fgf 4或Fgf 8的小鼠突变体，我们先前证明了它们冗余地阻止PSM分化。Fgf 8不是体节发生所必需的，但Fgf 4突变体显示出一系列椎骨缺陷。通过杂交链反应荧光标记的mRNA的基于空间模型的定量分析基因表达，我们表明FGF 4通过转录抑制因子HES 7控制Notch途径振荡。我们通过证明Hes 7和Fgf 4之间的遗传协同作用而不是与Fgf 8之间的遗传协同作用来支持这一假设。因此，我们建立Fgf 4作为一个必不可少的陷波振荡调节器和潜在的重要频谱的人类分割缺陷的椎骨缺陷所造成的缺陷陷波振荡。（eLife 2020年11月19日;9：e55608. doi：10.7554/eLife.55608.）未来的工作重点是Hes 7基因内的哪些调控元件对Fgf 4信号有反应。

To address the deficiency in our knowledge of what genes respond to FGF signaling, we have an ongoing project to molecularly define FGF targets genes, as well as their function and regulation. In past work, we and others have identified the transcription factors encoded by Gbx1 and Gbx2 as FGF targets. Recently we investigated the functional relationship between Gbx family members in the developing spinal cord using combinatorial Gbx mouse mutants. We showed that each Gbx gene is upregulated if the other is absent. Additionally, Gbx genes regulate development of a subset of PAX2+ dorsal inhibitory interneurons. Also, expansion of proliferative cells into the anatomically defined mantle zone occurs in Gbx mutants. Lastly, our data shows a marked increase in apoptotic cell death in the ventral spinal cord of Gbx mutants during mid-embryonic stages. While our studies reveal that both members of the Gbx gene family are involved in development of subsets of PAX2+ dorsal interneurons and survival of ventral motor neurons, Gbx1 and Gbx2 are not sufficient to genetically compensate for the loss of one another. Thus, our studies provide novel insight to the relationship harbored between Gbx1 and Gbx2 in spinal cord development (J Dev Biol. 2020. PMID: 32244588). In current work, we demonstrate that the Hes7 transcriptional repressor is apparently a direct target of Fgf4 signaling. During vertebrate development, the presomitic mesoderm (PSM) is periodically segmented into somites, which will form the segmented vertebral column and associated muscle, connective tissue, and dermis. The periodicity of somitogenesis is regulated by a segmentation clock of oscillating Notch activity. We examined mouse mutants lacking only Fgf4 or Fgf8, which we previously demonstrated act redundantly to prevent PSM differentiation. Fgf8 is not required for somitogenesis, but Fgf4 mutants display a range of vertebral defects. Analyzing gene expression with spatial model-based quantification of mRNAs fluorescently labeled by hybridization chain reaction, we show that FGF4 controls Notch pathway oscillations through the transcriptional repressor, HES7. We support this hypothesis by demonstrating a genetic synergy between Hes7 and Fgf4, but not with Fgf8. Thus, we establish Fgf4 as an essential Notch oscillation regulator and potentially important in a spectrum of human Segmentation Defects of the Vertebrae caused by defective Notch oscillations. (eLife 2020 Nov 19;9:e55608. doi: 10.7554/eLife.55608.) Future work focuses on what regulatory elements within the Hes7 gene are responsive to Fgf4 signals.