Regulatory Mechanisms Governing Vertebral Segmentation

椎骨分割的调节机制

• 批准号: 9316666
• 负责人: Ertugrul M Ozbudak
• 金额: $ 35.1万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9316666
• 关键词: Address; Affect; Binding; Binding Sites; Cell Differentiation process; Cell Proliferation; Cells; Circadian Rhythms; Congenital Abnormality; Defect; Elements; Embryonic Development; Feedback; Gene Expression; Gene Family; Genes; Genetic; Genetic Transcription; Heterogeneous-Nuclear Ribonucleoproteins; Human; Informatin; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Mathematics; Measures; Messenger RNA; Mission; Modeling; Mus; Mutation; Ovary; Pacemakers; Pathway interactions; Pattern; Periodicity; Process; Protein Family; Proteins; RNA Binding; RNA Decay; RNA-Binding Proteins; Role; Segmentation Clock Pathway; Stem cells; Testing; Time; Tissue Therapy; Tissues; Transcription Repressor/Corepressor; Ubiquitin; United States National Institutes of Health; Untranslated Regions; Vertebral column; Zebrafish; base; biological systems; cancer therapy; embryonic stem cell; experimental study; gain of function; gene discovery; in vivo; loss of function; mRNA Instability; malformation; multicatalytic endopeptidase complex; nerve stem cell; overexpression; predictive modeling; prevent; protein degradation; protein expression; public health relevance; spine bone structure; transcription factor; transcriptome sequencing; tumor progression

DESCRIPTION (provided by applicant): Gene expression oscillations are widespread in biological systems. The segmentation clock is one such oscillator controlling segmentation of the vertebral column. Its disruption results in congenital vertebral defects in humans. We have built a mathematical pacemaker model that is based on a transcriptional-feedback loop. Our model predicts that a moderate increase in the stabilities of Her proteins or mRNAs should lengthen the period, while a further increase would abolish the oscillations. To establish the mechanisms of Her protein and mRNA turnover, and to elucidate the mechanism that transfers this periodic information to cells of the next- forming segments, we will: 1. Discover the post-translational mechanism that rapidly recycles oscillating proteins: We will perform loss-of- function experiments for proteins that are candidates to regulate rapid degradation of Her proteins. We will assess whether any of these genes regulate the degradation of the Her-family proteins. Finally, to test the prediction of our model, we will stabilize Her proteins to varying levels by reducing expression of proteins that trigger its degradation and measure corresponding changes in the segmentation process. 2. Discover the post-transcriptional mechanism that rapidly recycles oscillating RNAs: We will determine loss- of-function of which her-RNA-binding proteins stabilize her mRNAs and result in vertebral segmentation defects. We will test the prediction of our model by increasing the halflives of her mRNAs through reduced expression of RNA-binding proteins and determining how the segmentation process is affected. 3. Discover the information transfer mechanism from the segmentation clock to the segmentation machinery: We will perform time-resolved overexpression and loss-of-function experiments for the mesp gene to determine its impact on the segmentation process. We will determine the regulatory cascade starting with the segmentation clock, continuing through the dynamically expressed mesp transcription factor and ending with the formation of segment boundaries. Hes proteins also oscillate in neural progenitor cells, ovary cells and embryonic stem cells, where the oscillations appear to control the temporal switch from proliferation to differentiation. Gain-of-function of Hes proteins is correlated with cancer and their inhibition restores differentiation. Elucidating the dynamics of the Hes/Her oscillations during vertebral segmentation is significant not only for understanding and potentially preventing vertebral malformations, but also for developing approaches for controlled stem cell proliferation and differentiation in various tissues and therapies against cancer progression. Therefore, this application has strong relevance to the mission of the National Institute of Health.

描述（由申请人提供）：基因表达振荡在生物系统中广泛存在。分段时钟是一种控制脊柱分段的振荡器。它的破坏会导致人类先天性脊椎缺陷。我们建立了一个基于转录反馈循环的数学起搏器模型。我们的模型预测，Her 蛋白或 mRNA 稳定性的适度增加会延长该周期，而进一步增加会消除振荡。为了建立 Her 蛋白和 mRNA 更新的机制，并阐明将这种周期性信息传递到下一个形成片段的细胞的机制，我们将： 1. 发现快速回收振荡蛋白的翻译后机制：我们将执行对作为调节 Her 蛋白快速降解的候选蛋白的功能丧失实验。我们将评估这些基因是否调节 Her 家族蛋白的降解。最后，为了测试我们模型的预测，我们将通过减少触发其降解的蛋白质表达来将 Her 蛋白质稳定到不同的水平，并测量分割过程中的相应变化。 2. 发现快速回收振荡RNA的转录后机制：我们将确定哪些her-RNA结合蛋白稳定她的mRNA并导致椎骨分段缺陷的功能丧失。我们将通过减少 RNA 结合蛋白的表达来增加 mRNA 的半衰期，并确定分割过程受到的影响，从而测试我们模型的预测。 3.发现从分割时钟到分割机器的信息传递机制：我们将对mesp基因进行时间分辨的过表达和功能丧失实验，以确定其对分割过程的影响。我们将确定从分段时钟开始的调控级联，继续通过动态表达的 mesp 转录因子，并以片段边界的形成结束。 Hes 蛋白也在神经祖细胞、卵巢细胞和胚胎干细胞中振荡，这些振荡似乎控制着从增殖到分化的时间转换。 Hes 蛋白的功能获得与癌症相关，其抑制可恢复分化。阐明椎体分割过程中 Hes/Her 振荡的动态不仅对于理解和潜在预防椎体畸形具有重要意义，而且对于开发各种组织中受控干细胞增殖和分化的方法以及对抗癌症进展的疗法也具有重要意义。因此，该应用程序与国立卫生研究院的使命密切相关。