Gene Regulatory Networks that Establish Mandible and Maxilla Patterning

建立下颌骨和上颌骨模式的基因调控网络

基本信息

批准号：
10454286
负责人：
David E. Clouthier
金额：
$ 59.85万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-01 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10454286
关键词：
Address Affect Alleles Area Automobile Driving BHLH Protein Bone structure Branchial arch structure CRISPR/Cas technology Cells Congenital Abnormality Craniofacial Abnormalities Data Defect Deformity Development Developmental Process Distant Dlx protein Dorsal Elements Environment Etiology Event Face First Pharyngeal Arch Future Gene Expression Gene Expression Profile Gene Targeting Genes Genetic Transcription Genetic study Goals Hand functions Health Helix-Turn-Helix Motifs Human Jaw Knowledge Live Birth Location Maintenance Mandible Mandibular Prominence Maps Maxillary Prominence Mediating Mediator of activation protein Mission Modeling Molecular Morphogenesis Mus Mutant Strains Mice Mutate Neural Crest Cell Operative Surgical Procedures Patients Pattern Pierre Robin Syndrome Play Process Public Health Quality of life Research Research Proposals Role Signal Transduction Smad Proteins Structural Genes Structure Syndrome Temporomandibular Joint Testing Transcriptional Regulation United States National Institutes of Health Zygomatic Arch craniofacial complex craniofacial development craniofacial structure dimer gain of function gene network gene regulatory network insight loss of function mandible/maxilla middle ear network models neural patterning novel novel strategies overexpression reconstruction single-cell RNA sequencing spatiotemporal synergism transcriptome

项目摘要

Abstract Craniofacial abnormalities affecting the mandible, maxilla and jaw joint are commonly encountered birth defects, most of which require surgical correction to establish quality of life and in some cases survival. While one or more organizing centers within the developing pharyngeal arches, from which the face arises, may hold the key to understanding the etiologies of these defects, the existence of such centers has never been proven, leading to a poor understanding of how gene regulatory networks are regulated and integrated during facial development. This limiting knowledge stifles new approaches to efficaciously treat these deformities. Endothelin1 (through DLX proteins) and BMP (through SMAD proteins) signaling establishes positional and structural identity of neural crest cells within the ventral mandibular arch. This is primarily achieved by induction of the basic bHLH transcription factor HAND2. HAND2 and BMP synergy is required for induction of the bHLH factor HAND1 within the ventral-most “cap” of the mandibular arch. Interestingly, altering HAND1 dimer partner choice (thus altering HAND1-mediated signaling) results in pronounced mid-facial clefting, even though Hand1 is not expressed in the mid-face structures. Our data shows that loss of either BMP or HAND2 activity disrupts the establishment of the ventral cap. These findings establish our hypothesis that the intersection of BMP and HAND2 activity establishes a ventral cap-signaling center, which acts in both cell and non-cell autonomous manners to drive upper and lower jaw development. Additionally, we hypothesize that DLX activity antagonizes BMP/HAND2 synergy. This proposal takes advantage of the craniofacial development expertise of Dr. David Clouthier, the bHLH signaling expertise of Dr. Anthony Firulli and a number of novel mutant mouse alleles to test these hypotheses in two Aims. In Aim 1, we will use single cell (sc) RNA-seq to define the gene regulatory networks that are initiated by the coordinated action of both HAND2 and BMP that act either in an autonomous (ventral cap) or non-cell autonomously (more dorsal first arch areas) manner. Following analysis of scRNA-seq data, the top HAND2/BMP effector candidates will be functionally evaluated in loss-of-function and genetic studies. In Aim 2, the role of DLX proteins in confining ventral cap size will be examined using a novel gain-of- function Dlx5 mouse allelefollowed by functional testing of DLX action by creating a Hand1 mouse mutant lacking DLX cis-element inputs. Together, these novel approaches will provide the first direct evidence that the mandibular arch ventral cap is a signaling center required for facial development. Relevance: Craniofacial abnormalities are common and require intensive reconstructive surgical corrections. HAND2 and BMPs play key roles in patterning the neural crest cells that form the face. Gaining insight into the molecular mechanism of this understudied developmental process could have great potential for initial development of non-surgical treatments for congenital craniofacial defects in patients.

抽象的颅面异常影响下颌骨，上颌骨和下颌关节通常遇到出生缺陷，其中大多数需要手术校正才能确定生活质量，在某些情况下可以生存。尽管发育中的咽弓内的一个或多个组织中心，面部出现的一个或了解这些缺陷的病因的关键，从未证明此类中心的存在，导致对基因调节网络如何调节和整合在面部期间的不良理解发展。这种限制知识扼杀了有效治疗这些畸形的新方法。内皮素1（通过DLX蛋白质）和BMP（通过SMAD蛋白）信号传导建立位置和腹侧下颌弓内神经rest细胞的结构认同。这主要是通过归纳来实现的基本BHLH转录因子手2。 Hand2和BMP协同作用是BHLH的诱导所必需的因子hand1在下颌拱的最腹侧“盖”中。有趣的是，更改Hand1 Dimer合作伙伴选择（从而改变了手动1介导的信号传导）也会导致明显的中间裂纹，即使Hand1 在中面结构中不表示。我们的数据表明，BMP或Hand2活动的损失会破坏建立腹帽。这些发现确定了我们的假设，即BMP与 Hand2活性建立了一个腹侧帽信号中心，该中心在细胞和非电池自动群中起作用驱动上下颌开发的举止。另外，我们假设DLX活性拮 BMP/Hand2协同作用。该建议利用了大卫博士的颅面发展专家 Clouthier，Anthony Firulli博士的BHLH信号专业知识以及许多新型的突变小鼠等位基因在两个目标中检验这些假设。在AIM 1中，我们将使用单细胞（SC）RNA-Seq来定义基因调节由Hand2和BMP的协调动作发起的网络，它们要么在自主中行动（腹盖）或非单元自主（更多背弓区域）的方式。以下分析scrna-seq 数据，顶部2/BMP效应子候选者将在功能丧失和遗传方面进行功能评估研究。在AIM 2中，DLX蛋白在限制通风帽尺寸中的作用将使用新颖的收益来检查功能DLX5鼠标通过创建Hand1 Mouse突变体的DLX动作的功能测试来遵循函数。缺少DLX顺元元素输入。这些新颖的方法一起将提供第一个直接证据下颌拱腹盖是面部发育所需的信号中心。相关性：颅面异常是常见的，需要进行密集的重建手术校正。 HAND2和BMP在对形成面部的神经rest细胞的图案中起关键作用。洞悉这种理解的发展过程的分子机制可能具有很大的初始潜力开发患者先天性颅面缺陷的非手术治疗。