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：
10057669
负责人：
David E. Clouthier
金额：
$ 62.38万
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/10057669
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 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 Regulator Genes 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 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.

抽象的影响下颌骨、上颌骨和颌关节的颅面畸形是出生时常见的情况缺陷，其中大多数需要手术矫正才能提高生活质量，在某些情况下还需要生存。发育中的咽弓内的一个或多个组织中心（面部从咽弓中产生）可能包含了解这些缺陷病因的关键是，此类中心的存在从未得到证实，导致人们对面部护理过程中基因调控网络的调控和整合方式缺乏了解这种有限的知识阻碍了有效治疗这些畸形的新方法。内皮素1（通过DLX蛋白）和BMP（通过SMAD蛋白）信号传导建立位置和腹侧下颌弓内神经嵴细胞的结构特征这主要是通过诱导来实现的。 bHLH 的诱导需要基本 bHLH 转录因子 HAND2 和 BMP 的协同作用。下颌弓最腹侧“帽”内的因子 HAND1。选择（从而改变 HAND1 介导的信号传导）会导致明显的中面部裂，即使 Hand1 我们的数据表明，BMP 或 HAND2 活性的丧失会破坏中面部结构。这些发现证实了我们的假设，即 BMP 和腹侧盖的交叉点。 HAND2 活动建立腹侧帽信号传导中心，该中心在细胞和非细胞自主中发挥作用促进上颌和下颌发育的方式。 BMP/HAND2 协同作用该提案利用了 David 博士的颅面发育专业知识。 Clouthier、Anthony Firulli 博士的 bHLH 信号转导专业知识和许多新型突变小鼠等位基因在两个目标中测试这些假设。在目标 1 中，我们将使用单细胞 (sc) RNA-seq 来定义基因调控。网络是由 HAND2 和 BMP 的协调行动发起的，它们要么以自主的方式行动，要么以自主的方式行动。（腹帽）或非细胞自主（更多背侧第一弓区域）方式进行 scRNA-seq 分析。数据显示，顶级 HAND2/BMP 候选效应器将在功能丧失和遗传方面进行功能评估在目标 2 中，将使用一种新的增益来检查 DLX 蛋白在限制腹帽大小中的作用。功能 Dlx5 小鼠等位基因，然后通过创建 Hand1 小鼠突变体对 DLX 作用进行功能测试缺乏 DLX 顺式元件输入，这些新方法将提供第一个直接证据。下颌弓腹帽是面部发育所需的信号中心。相关性：颅面异常很常见，需要密集的重建手术矫正。 HAND2 和 BMP 在形成面部神经嵴细胞的模式中发挥着关键作用。这种正在研究的发育过程的分子机制可能具有巨大的初始潜力开发针对先天性颅面缺陷患者的非手术治疗方法。