Determining the role of sema3d during fin regeneration

确定 sema3d 在鳍再生过程中的作用

• 批准号: 8446293
• 负责人: Mary Kathryn Iovine
• 金额: $ 6.96万
• 依托单位: LEHIGH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2014-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8446293
• 关键词: Affect; Axon; Biological Assay; Biological Models; Blood Vessels; Bone Growth; Cell Differentiation process; Cell Proliferation; Cell Proliferation Regulation; Cells; Connexin 43; Cues; Data; Defect; Development; Distal; Event; Exhibits; Failure; Gap Junctions; Genes; Goals; Growth; Human; Joints; Lead; Length; Limb structure; Location; Mediating; Molecular; Morphogenesis; Morphology; Multiple Sclerosis; Mutation; Natural regeneration; Neurodegenerative Disorders; Outcome; Pathway interactions; Pattern; Phenotype; Rheumatoid Arthritis; Role; Semaphorins; Shapes; Signal Pathway; Signal Transduction; Signaling Molecule; Skeleton; Syndrome; Tissues; Zebrafish; cell type; craniofacial; gain of function; human disease; insight; loss of function; member; mutant; oculodentodigital dysplasia; osteoblast differentiation; premature; receptor; skeletal; tumor progression

DESCRIPTION (provided by applicant): Skeletal morphogenesis requires the integration of multiple signals for coordinated growth and patterning. For example, cell proliferation must be coordinated with the differentiation of osteoblasts and joint-forming cells. However, the mechanisms responsible for coordinating multiple cellular events are largely unknown. Our analyses using the zebrafish regenerating fin as a model system have identified two fin length mutants that provide insight into the coordination of fin growth and skeletal patterning. The short fin (sof b123) mutant has short fins due to the development of short fin ray segments (i.e. or premature joint formation). In contrast, the another long fin (alf dty86) mutant exhibits fin overgrowth and overlong segments due to stochastic joint failure. Interestingly, sof b123and alf dty86 also exhibit important and opposing differences in the expression of the gap junction gene connexin43 (cx43). The sof b123phenotypes are caused by hypomorphic mutations in cx43, while the joint failure phenotype in alf dty86 is the result of cx43 over-expression. Indeed, cx43 knockdown in alf dty86regenerating fins rescues joint formation. Together, these and other data suggest that Cx43 function both promotes fin growth and suppresses joint formation. Moreover, mutations in human CX43 cause oculodentodigital dysplasia, a syndrome characterized by morphological defects of the craniofacial and distal limb skeletons. Thus, Cx43 exhibits conserved, but unknown, functions in skeletal morphogenesis. In order to understand molecularly how Cx43 mediates its effects on growth and patterning of the fin skeleton, we identified genes expressed downstream of cx43. Interestingly, the gene for a secreted semaphorin, sema3d, is expressed in the regenerating fin in a cx43-dependent manner. Strikingly, knockdown of sema3d leads to reduced fin length, reduced cell proliferation, reduced segment length, and rescue of joint formation in alf dty86, similar to the cx43-knockdown phenotypes. Together, our findings suggest that sema3d functions in a common pathway with cx43 to regulate fin growth and joint formation. Semaphorins are members of a large evolutionarily conserved class of signaling molecules, best understood as providing guidance cues for axons and blood vessels. More recent studies revealed that semaphorins are expressed broadly during development and initiate a wide diversity of cellular outcomes. Loss of semaphorin function leads to human diseases including cancer progression, neurodegenerative diseases, multiple sclerosis, and rheumatoid arthritis. Thus, semaphorins contribute broadly to normal and pathological development. The aims of this proposal are to identify the cx43-dependent cellular outcomes affected by manipulating Sema3d signaling and to identify the components of the signaling pathway downstream of Sema3d. Results obtained from this proposal will reveal the underlying mechanism of Cx43-Sema3d action during fin regeneration, and will serve as preliminary data for a more comprehensive proposal on defining the functional relationship between Cx43 and Sema3d, and the molecular pathway(s) required for the coordination of growth and patterning of the vertebrate skeleton.

描述（由申请人提供）：骨骼形态发生需要将多个信号集成以进行协调的生长和模式。例如，细胞增殖必须与成骨细胞和关节形成细胞的分化协调。但是，负责协调多个细胞事件的机制在很大程度上未知。我们使用斑马鱼再生鳍作为模型系统的分析已经确定了两个鳍长的突变体，这些突变体提供了对鳍片生长和骨骼模式的配位的见解。短 由于短鳍射线段（即或过早的关节形成），FIN（SOF B123）突变体具有短鳍。相比之下，另一个长鳍（ALF DTY86）突变体表现出由于随机关节失败而导致的鳍过度生长和延长的细分市场。有趣的是，SOF B123和ALF DTY86在间隙连接基因connexin43（CX43）的表达上也表现出重要和相反的差异。 SOF B123PheNotypes是由CX43中的型突变引起的，而ALF DTY86中的关节衰竭表型是CX43过表达的结果。实际上，ALF DTY86的鳍片中的CX43敲低鳍挽救了联合形成。这些数据和其他数据一起表明CX43功能既可以促进鳍的生长并抑制关节形成。此外，人CX43中的突变引起眼缘牙性牙本质增生，这是一种以颅面和远端肢体骨骼形态缺陷为特征的综合征。因此，CX43在骨骼形态发生中表现出保守但未知的功能。为了了解CX43如何介导其对鳍骨架的生长和模式的影响，我们确定了CX43下游表达的基因。有趣的是，分泌的信号素Sema3d的基因以CX43依赖性方式在再生鳍中表达。令人惊讶的是，SEMA3D的敲低导致鳍长度降低，细胞增殖减少，段长度减少以及与ALF DTY86中关节形成的拯救，类似于CX43-KNOCKDOWN STYPES。总之，我们的发现表明SEMA3D在具有CX43的共同途径中起作用，以调节鳍的生长和关节形成。信号素是大型进化保守的信号分子类别的成员，最好理解为为轴突和血管提供指导线索。最近的研究表明，闪光蛋白在发育过程中广泛表达，并引发了各种细胞结局。信号素功能的丧失导致人类疾病，包括癌症进展，神经退行性疾病，多发性硬化症和类风湿关节炎。因此，信号素对正常和病理发育有广泛的贡献。该提案的目的是确定受SEMA3D信号传导影响的CX43依赖性细胞结果，并确定SEMA3D下游信号通路的组件。从该提案中获得的结果将揭示FIN再生过程中CX43-SEMA3D动作的潜在机制，并将作为定义CX43和SEMA3D之间功能关系的更全面的建议，以及分子途径的更全面的建议，以及用于生长和统治验证的分子途径（S）。