Hoxd and Gli3-Hoxd interaction roles in Hedgehog regulated digit morphogenesis

Hoxd 和 Gli3-Hoxd 相互作用在 Hedgehog 调节数字形态发生中的作用

• 批准号: 10702480
• 负责人: Susan Mackem
• 金额: $ 54.4万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10702480
• 关键词: Adult; Ambystoma; Amphibia; Anterior; Appearance; Cartilage; Cell Adhesion; Cell Aggregation; Cell Death; Cell physiology; Cessation of life; Chromatin Remodeling Factor; Complex; Congenital Abnormality; Development; Developmental Biology; Digit structure; Disease; Distal; Elements; Ensure; Equilibrium; Erinaceidae; Evolution; Excision; Failure; Feedback; Fibroblast Growth Factor; Finger joint structure; Fingers; GLI3 gene; Gene Dosage; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genomics; Goals; Growth; Homeobox; Homeostasis; Human; Intercept; Joints; Lead; Learning; Length; Limb Bud; Limb Development; Limb structure; Link; Maintenance; Malignant Neoplasms; Mammals; Mesenchyme; Mission; Morphogenesis; Morphology; Mus; Natural regeneration; Neoplasm Metastasis; Neoplasms; Organ; Output; Pathologic; Pathology; Pathway interactions; Pattern; Periodicity; Phalanx; Phalanx of hand; Phase; Physical condensation; Physiological Processes; Play; Positioning Attribute; Process; Proteins; Proteomics; Radial; Regenerative capacity; Regulation; Replication Licensing; Research; Role; Running; SHH gene; Shapes; Signal Transduction; Site; Skeleton; Skin; Structure; Study models; System; Systems Biology; Thumb structure; Tissues; Tumor Biology; Vertebrates; Work; austin; base; beta catenin; cell behavior; cell motility; cellular targeting; design; gastrointestinal epithelium; genetic approach; insight; joint formation; mutant; neoplastic; neoplastic cell; progenitor; programs; regeneration potential; selective expression; skeletal; skeletal regeneration; smoothened signaling pathway; transcription factor; tumorigenesis; ulna

5'Hoxd genes play many roles during limb development and may control the effectors of morphogenesis at late stages. How Hoxd genes guide digit morphogenesis and their downstream targets remain enigmatic. Using genetic approaches in mice we find that, in addition to a role in initiating Sonic hedgehog (Shh) expression, 5'Hoxd genes determine the polarity of the primary limb axis early, and regulate digit pattern and morphogenesis at late stages, after digit condensations have already formed, including joint formation and positioning; a major mechanism by which Hoxd genes regulate digit identity. We previously discovered genetic and physical interactions between 5Hoxd and Gli3 that modify Gli3 repressor (Gli3R) function (and hence Shh output), antagonizing Gli3R and potentially converting it to an activator. We find that Gli3-Hox interactions both modulate the polarity of limb axis formation and regulate the pacing of cartilage vs joint formation in digits, which may have relevance for skeletal homeostasis and disease, as well as skeletal birth defects. ....................................................................................................................................................................................................................................................................................................................Role Role of 5'Hoxd genes and Hoxd-Gli3 interaction in determining polarity of primary limb axis formation: In most vertebrates, the primary limb axis runs through the posterior limb with the ulna/digit4 (d4) condensing first. In urodele amphibians such as axolotl, which retain the ability to regenerate limbs as adults, the anterior limb axis is dominant (radius/d2 appear first). Based on altered expression patterns, it has been proposed that the axis shift in Urodeles results from a failure to expand 5'Hoxd gene expression in the late distal limb. We have analyzed limb axis formation in the 5'Hoxd mutant (Hoxd11-13 deleted) and found that the anterior axis forms first as in urodeles. Furthermore, we find that in compound 5'Hoxd;Gli3 mutants, posterior axial dominance is restored. The 5'Hoxd homeobox transcription factors play roles in replication licensing and cell adhesion. Gli3R, expressed anteriorly, also regulates proliferation and condensation, and antagonizes 5'Hoxd function. We are analyzing how changes in the relative timing and rate of proliferation and of cell aggregation/condensation in different zones of the limb bud are altered in these mutants, and if they correlate with anterior vs posterior axial dominance. We propose that the balance between antagonistic 5'Hoxd-Gli3 functions governs the polarity of primary limb axis formation and are investigating the potential relation between altered axis polarity and regenerative capacity. ....................................................................................................................................................................................................................................................................................................................Role of Hoxd genes and Hoxd-Gli3 interaction in cartilage differentiation and joint formation to determine distinct digit identities: Digit identity remains plastic even after the formation of the digit primordial chondrogenic condensations and is regulated by interdigit zones, which are also late sites of 5'Hoxd and Gli3 expression. We found that genetic removal of several Hoxd genes (d11-d13) results in abnormal joint formation, both loss of digit joints and/or abnormal joint position, as well as short, biphalangeal digits. Collaborating with Marian Ros (Univ. Cantabria) we are also examining the role of Hoxa13 in digit formation. Hoxa13 acts upstream of and induces the late phase of Hoxd13 expression and plays a distinct role in regulating the formation of a normal thumb acting hierarchically upstream of, and together with Hoxd13. The canonical Wnt pathway plays an essential role in joint formation and we find that activated beta-catenin restores normal joint formation in the 5'Hoxd mutant digits. But surprisingly, selective activation of stabilized beta-catenin in the interdigital tissues is required for rescue, indicating that at least some aspects of beta-catenin and 5'Hoxd function in joint formation occur indirectly, via interdigit signaling. Gli3 (the transcriptional effector of Shh and Hoxd protein interactor) also has striking effects on cartilage differentiation and joint formation in digits. During joint formation in digit precursors, Gli3 mutants form abnormal segments with excessive joint formation extending into the cartilage elements. Genetically, the balance between total 5'Hoxd and Gli3 gene dosage regulates the periodic formation of normal joints and the normal 3 bony segments typical of mammalian digits. Our genetic evidence indicates that the Hoxd-Gli3 balance acts indirectly, from interdigital mesenchyme, to modulate Bmp activity and thereby regulate the periodic appearance of digit elements (phalanges) and joints from a digit tip progenitor pool. We are extending our analysis to determine: 1) targets regulated by Gli3-Hoxd interaction and 2) other signaling inputs that regulate the digit tip progenitor pool to determine phalanx number and size, including signals induced by beta-catenin activation in interdigits. Collaborating with Steve Vokes (UT-Austin) we will examine the role of the chromatin modifier Prmt5, which is selectively expressed in the digit tip progenitors, in maintaining this progenitor pool. Even in mammals, distal digit tips retain a limited capacity for regeneration and understanding the regulation of this distal digit progenitor pool and it's maintenance will provide new insights relevant to skeletal regeneration potential.

5'Hoxd基因在肢体发育过程中发挥了许多作用，并可能在后期控制形态发生的效应。Hoxd基因如何引导手指形态发生及其下游目标仍然是一个谜。通过对小鼠的遗传学研究，我们发现除了启动Sonic hedgehog （Shh）表达外，5'Hoxd基因在早期决定主要肢体轴的极性，并在手指缩合形成后的后期调节手指模式和形态发生，包括关节的形成和定位；这是Hoxd基因调控数字身份的主要机制。我们之前发现了5Hoxd和Gli3之间的遗传和物理相互作用，可以改变Gli3抑制因子（Gli3R）的功能（以及Shh的输出），拮抗Gli3R并可能将其转化为激活因子。我们发现Gli3-Hox相互作用既可以调节肢轴形成的极性，也可以调节手指软骨与关节形成的节奏，这可能与骨骼稳态和疾病有关。以及骨骼出生缺陷 . ....................................................................................................................................................................................................................................................................................................................5'Hoxd基因和Hoxd-Gli3相互作用在决定主肢轴形成极性中的作用：在大多数脊椎动物中，主肢轴穿过后肢，尺骨/指（d4）首先凝结。在蝾螈等尾纲两栖动物中，它们在成年后仍具有肢体再生的能力，前肢轴占主导地位（首先出现半径/d2）。基于表达模式的改变，有人提出尾犬的轴移是由于末端肢体中5'Hoxd基因表达扩展失败造成的。我们分析了5'Hoxd突变体（Hoxd11-13缺失）的肢轴形成，发现与尾犬一样，前肢轴首先形成。此外，我们发现化合物5'Hoxd；Gli3突变体，后轴优势恢复。5'Hoxd同源盒转录因子在复制许可和细胞粘附中起作用。Gli3R，表达在前面，也调节增殖和缩聚，并拮抗5'Hoxd功能。我们正在分析在这些突变体中，肢体芽不同区域的增殖和细胞聚集/凝结的相对时间和速率的变化是如何改变的，以及它们是否与前轴优势和后轴优势相关。我们提出拮抗5'Hoxd-Gli3功能之间的平衡决定了主肢轴形成的极性，并正在研究轴极性改变与再生能力之间的潜在关系。....................................................................................................................................................................................................................................................................................................................Hoxd基因和Hoxd-Gli3相互作用在软骨分化和关节形成中确定不同手指身份的作用：即使在手指原始软骨凝聚形成后，手指身份仍然具有可塑性，并受指间区调节，趾间区也是5'Hoxd和Gli3表达的后期位点。我们发现几个Hoxd基因（d11-d13）的遗传去除会导致关节形成异常，手指关节丢失和/或关节位置异常，以及短的双指指。与Marian Ros（坎塔布里亚大学）合作，我们也在研究Hoxa13在手指形成中的作用。Hoxa13作用于Hoxd13表达的上游，并诱导Hoxd13的后期表达，在调节正常拇指的形成中发挥明显作用，并与Hoxd13一起作用于Hoxd13的上游。典型的Wnt通路在关节形成中起着至关重要的作用，我们发现激活的β -连环蛋白可以恢复5'Hoxd突变趾的正常关节形成。但令人惊讶的是，在指间组织中，稳定的β -catenin的选择性激活是修复所必需的，这表明β -catenin和5'Hoxd在关节形成中的作用至少在某些方面是通过指间信号间接发生的。Gli3 （Shh和Hoxd蛋白相互作用因子的转录效应因子）在趾软骨分化和关节形成中也有显著的影响。在手指前体的关节形成过程中，Gli3突变体形成异常节段，过度的关节形成延伸到软骨元件。从遗传学上讲，5'Hoxd和Gli3基因总剂量之间的平衡调节着正常关节和哺乳动物指骨典型的正常3骨节的周期性形成。我们的遗传证据表明，Hoxd-Gli3平衡间接地通过指间间质调节Bmp活性，从而调节指尖祖细胞池中指元素（指骨）和关节的周期性出现。我们正在扩展我们的分析，以确定：1)Gli3-Hoxd相互作用调节的靶标和2)调节指尖祖细胞池的其他信号输入，以确定指骨数目和大小，包括指间β -连环蛋白激活诱导的信号。与Steve Vokes （UT-Austin）合作，我们将研究在指尖祖细胞中选择性表达的染色质修饰子Prmt5在维持这个祖细胞库中的作用。即使在哺乳动物中，远端趾尖也保留了有限的再生能力，了解远端趾祖池的调节及其维护将为骨骼再生潜力提供新的见解。