Regulatory Mechanisms Governing Vertebral Segmentation

椎骨分割的调节机制

• 批准号: 8930167
• 负责人: Ertugrul M Ozbudak
• 金额: $ 3.13万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-20 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8930167
• 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; Health; Heterogeneous-Nuclear Ribonucleoproteins; Human; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Measures; Messenger RNA; Mission; Modeling; Mus; Mutation; Ovary; Pacemakers; Pathway interactions; Pattern; Process; Protein Family; Proteins; RNA Decay; RNA Sequences; RNA-Binding Proteins; Recycling; Role; Segmentation Clock Pathway; Stem cells; Testing; Time; Tissue Therapy; Tissues; Transcription Repressor/Corepressor; Transcription factor genes; Ubiquitin; United States National Institutes of Health; Untranslated Regions; Vertebral column; Zebrafish; base; biological systems; cancer therapy; embryonic stem cell; gain of function; gene discovery; in vivo; loss of function; mRNA Instability; malformation; multicatalytic endopeptidase complex; nerve stem cell; overexpression; prevent; protein expression; research study; 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。分段时钟是控制脊柱分段的一个这样的振荡器。它的破坏导致人类先天性脊椎缺陷。我们建立了一个基于转录反馈回路的数学起搏器模型。我们的模型预测，Her蛋白质或mRNA的稳定性的适度增加应该会延长周期，而进一步增加则会消除振荡。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给下一个形成节段的细胞的机制，我们将：1。发现快速降解振荡蛋白的翻译后机制：我们将对调节Her蛋白快速降解的候选蛋白进行功能丧失实验。我们将评估这些基因中是否有任何一个调节Her家族蛋白质的降解。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的表达来稳定Her蛋白质到不同的水平，并测量分割过程中的相应变化。2.发现转录后机制，迅速使振荡的RNA：我们将确定功能丧失的her-RNA结合蛋白稳定她的mRNA，并导致椎骨分割缺陷。我们将通过减少RNA结合蛋白的表达来增加她的mRNA的半衰期，并确定分割过程如何受到影响，来测试我们模型的预测。3.发现从分割时钟到分割机制的信息传递机制：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节级联，继续通过动态表达的mesp转录因子，并以分段边界的形成结束。 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，并且它们的抑制恢复分化。阐明在椎骨分割过程中Hes/Her振荡的动力学不仅对于理解和潜在地预防椎骨畸形，而且对于开发用于在各种组织中控制干细胞增殖和分化的方法和针对癌症进展的疗法是重要的。因此，本申请与国家卫生研究所的使命具有很强的相关性。