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: 10014540
• 负责人: Susan Mackem
• 金额: $ 59.96万
• 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/10014540
• 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; 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; digit regeneration; gastrointestinal epithelium; genetic approach; insight; joint formation; mutant; neoplastic; neoplastic cell; progenitor; programs; regenerative; 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 may have a distinct role in regulating the formation of a normal thumb. 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, suggesting 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. 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多种作用，并可能控制后期形态发生的效应器。 Hoxd 基因如何指导数字形态发生及其下游目标仍然是个谜。通过在小鼠中使用遗传方法，我们发现，除了在启动音刺猬 (Shh) 表达中发挥作用外，5'Hoxd 基因还早期确定主肢轴的极性，并在手指压缩后的后期调节手指模式和形态发生已经形成，包括接头形成和定位； Hoxd 基因调节数字身份的主要机制。我们之前发现 5Hoxd 和 Gli3 之间的遗传和物理相互作用会改变 Gli3 阻遏蛋白 (Gli3R) 功能（从而产生 Shh 输出），拮抗 Gli3R 并可能将其转化为激活剂。我们发现 Gli3-Hox 相互作用既能调节肢体轴形成的极性，又能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以及骨骼出生缺陷有关。 ...................................................... ...................................................... ...................................................... ...................................................... ...................................................... ...................................................... ........作用 5'Hoxd 基因和 Hoxd-Gli3 相互作用在决定初级肢体轴形成的极性中的作用：在大多数脊椎动物中，初级肢轴通过尺骨/数字4（d4）穿过后肢先凝结。在有尾目两栖动物（例如蝾螈）中，成年后仍保留肢体再生能力，前肢轴占主导地位（半径/d2首先出现）。基于表达模式的改变，有人提出，有尾目的轴移是由于晚期远端肢体中 5'Hoxd 基因表达未能扩展所致。我们分析了 5'Hoxd 突变体（Hoxd11-13 删除）中肢轴的形成，发现前轴首先形成，就像在无尾动物中一样。此外，我们发现在化合物 5'Hoxd;Gli3 突变体中，后轴优势得以恢复。 5'Hoxd 同源框转录因子在复制许可和细胞粘附中发挥作用。前面表达的 Gli3R 还调节增殖和凝结，并拮抗 5'Hoxd 功能。我们正在分析这些突变体中肢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如何改变，以及它们是否与前轴优势和后轴优势相关。我们提出拮抗性 5'Hoxd-Gli3 功能之间的平衡控制初级肢体轴形成的极性，并正在研究改变的轴极性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的作用，以确定不同的数字身份：即使在数字原始软骨形成凝结形成后，数字身份仍然保持可塑性，并受到指间区域的调节，它们也是 5'Hoxd 和 Gli3 表达的晚期位点。我们发现，几个 Hoxd 基因 (d11-d13) 的基因去除会导致关节形成异常、手指关节缺失和/或关节位置异常，以及双指手指短小。我们还与 Marian Ros（坎塔布里亚大学）合作研究 Hoxa13 在数字形成中的作用。 Hoxa13 在 Hoxd13 表达的上游起作用并诱导其后期表达，并且可能在调节正常拇指的形成中具有独特的作用。经典的 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-catenin 可以恢复 5'Hoxd 突变手指的正常关节形成。但令人惊讶的是，拯救需要选择性激活指间组织中稳定的 β-连环蛋白，这表明 β-连环蛋白和 5'Hoxd 在关节形成中的功能至少在某些方面是通过指间信号传导间接发生的。 Gli3（Shh 和 Hoxd 蛋白相互作用子的转录效应子）对手指软骨分化和关节形成也具有显着影响。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中。从遗传学上来说，5'Hoxd 和 Gli3 基因总剂量之间的平衡调节正常关节和哺乳动物手指典型的正常 3 个骨节的周期性形成。我们的遗传证据表明，Hoxd-Gli3 平衡通过指间充质间接发挥作用，调节 Bmp 活性，从而调节指尖祖细胞库中指骨（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）受 Gli3-Hoxd 相互作用调节的目标，以及 2）调节指尖祖细胞库以确定指骨数量和大小的其他信号输入。我们将与 Steve Vokes（UT-Austin）合作，研究染色质修饰剂 Prmt5 在维持该祖细胞库中的作用，Prmt5 在指尖祖细胞中选择性表达。即使在哺乳动物中，远端指尖也保留有限的再生能力，了解远端指祖细胞池的调节及其维护将为骨骼再生潜力提供新的见解。