Dysferlin regulation of acetylcholine signaling at the C. elegans NMJ

Dysferlin 对线虫 NMJ 乙酰胆碱信号传导的调节

• 批准号: 8000546
• 负责人: Jessica E Tanis
• 金额: $ 5.05万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8000546
• 关键词: Acetylcholine; Affect; Agonist; Animal Model; Animals; Behavioral Assay; Caenorhabditis elegans; Cells; Cholinergic Receptors; DYSF gene; Defect; Development; Event; Exhibits; Functional disorder; Genes; Genetic; Hereditary Disease; Lead; Levamisole; Limb-Girdle Muscular Dystrophies; Locomotion; Mediating; Membrane; Modeling; Molecular; Muscle; Muscle Weakness; Muscle function; Muscular Dystrophies; Mutation; Myopathy; Neuromuscular Junction; Neurons; Orthologous Gene; Phenotype; Play; RNA Interference; Regulation; Research; Resistance; Role; Signal Transduction; Site; Skeletal Muscle; Synapses; System; Testing; Therapeutic; United States; Vesicle; in vivo; loss of function mutation; mutant; novel; promoter; public health relevance; repaired; research study; synaptic function

DESCRIPTION (provided by applicant): The muscular dystrophies, incurable genetic disorders that result in progressive muscle weakness and degeneration, affect about 250,000 people in the United States. Genetic causes of many muscular dystrophies are known, however, further understanding of muscular dystrophy pathophysiology is required to develop appropriate therapeutic strategies. Limb Girdle Muscular Dystrophy type 2B (LGMD2B) is caused by loss of function mutations in Dysferlin, which regulates vesicle fusion events to repair damaged muscle membranes. Exactly how loss of Dysferlin lead to LGMD2B phenotypes is unknown. Exciting results in C. elegans show that the Dysferlin ortholog fer-1 is expressed in body-wall muscles where it plays a novel role in synaptic function by regulating the localization of acetylcholine receptors (AChRs) at the neuromuscular junction (NMJ). Integration of these findings with previous models of Dysferlin- mediated vesicle fusion, suggests that loss of FER-1/Dysferlin causes a reduction in AChR-containing vesicle fusion at the NMJ, leading to defects in synaptic function that may contribute to LGMD2B phenotypes. Using the model organism C. elegans, three independent lines of experimentation will be used to study this novel synaptic role of FER-1 and more broadly, the regulation of post-synaptic acetylcholine (ACh) signaling. C. elegans is a powerful genetic system used for analysis of muscle function, and the body-wall muscles which control C. elegans locomotion are functionally comparable to vertebrate skeletal muscle. Although C. elegans fer-1 is expressed in muscles, its site of action is not known. Cell-specific promoters will be used to express fer-1 in muscles or neurons in order to determine where FER-1 functions. Additional rescue experiments will be performed to determine if C. elegans fer-1 and mammalian Dysferlin are functionally orthologous in the regulation of synaptic function. Although fer-1 mutants exhibit defects in pharmacological behavioral assays and the localization of post-synaptic AChRs, the effect of loss of fer-1 on body-wall muscle activity is unknown. Thus, an in vivo electrophysiological approach will be used to determine the effect of fer-1 mutations on acetylcholine- evoked muscle currents and further define the role of C. elegans FER-1 in synaptic function. Finally, the molecular mechanisms that regulate and maintain proper post-synaptic acetylcholine (ACh) signaling are not fully understood. Additional genes that, like fer-1, are required for the modulation of ACh signaling will be identified by performing an RNA interference (RNAi) screen for animals resistant to the AChR agonist levamisole. In conclusion, I will achieve a further understanding of the molecular mechanisms underlying ACh signaling at the NMJ by testing a novel role for FER-1/Dysferlin in the regulation of synaptic function and identifying novel genes required for regulation of post-synaptic ACh signaling. PUBLIC HEALTH RELEVANCE: Limb Girdle Muscular Dystrophy type 2B (LGMD2B), which is caused by loss of function mutations in the gene Dysferlin, results in progressive muscle weakness. Recent results suggest that loss of Dysferlin causes defects in synaptic function and this may contribute to LGMD2B phenotypes. Our research using C. elegans to study this novel pathophysiological mechanism of LGMD2B may lead to a better understanding of the molecular mechanisms that cause progressive muscle weakness and could lead to the development of new therapies to treat this incurable muscle disease.

DESCRIPTION (provided by applicant): The muscular dystrophies, incurable genetic disorders that result in progressive muscle weakness and degeneration, affect about 250,000 people in the United States. Genetic causes of many muscular dystrophies are known, however, further understanding of muscular dystrophy pathophysiology is required to develop appropriate therapeutic strategies. Limb Girdle Muscular Dystrophy type 2B (LGMD2B) is caused by loss of function mutations in Dysferlin, which regulates vesicle fusion events to repair damaged muscle membranes. Exactly how loss of Dysferlin lead to LGMD2B phenotypes is unknown. Exciting results in C. elegans show that the Dysferlin ortholog fer-1 is expressed in body-wall muscles where it plays a novel role in synaptic function by regulating the localization of acetylcholine receptors (AChRs) at the neuromuscular junction (NMJ). Integration of these findings with previous models of Dysferlin- mediated vesicle fusion, suggests that loss of FER-1/Dysferlin causes a reduction in AChR-containing vesicle fusion at the NMJ, leading to defects in synaptic function that may contribute to LGMD2B phenotypes. Using the model organism C. elegans, three independent lines of experimentation will be used to study this novel synaptic role of FER-1 and more broadly, the regulation of post-synaptic acetylcholine (ACh) signaling. C. elegans is a powerful genetic system used for analysis of muscle function, and the body-wall muscles which control C. elegans locomotion are functionally comparable to vertebrate skeletal muscle. Although C. elegans fer-1 is expressed in muscles, its site of action is not known. Cell-specific promoters will be used to express fer-1 in muscles or neurons in order to determine where FER-1 functions. Additional rescue experiments will be performed to determine if C. elegans fer-1 and mammalian Dysferlin are functionally orthologous in the regulation of synaptic function. Although fer-1 mutants exhibit defects in pharmacological behavioral assays and the localization of post-synaptic AChRs, the effect of loss of fer-1 on body-wall muscle activity is unknown. Thus, an in vivo electrophysiological approach will be used to determine the effect of fer-1 mutations on acetylcholine- evoked muscle currents and further define the role of C. elegans FER-1 in synaptic function. Finally, the molecular mechanisms that regulate and maintain proper post-synaptic acetylcholine (ACh) signaling are not fully understood. Additional genes that, like fer-1, are required for the modulation of ACh signaling will be identified by performing an RNA interference (RNAi) screen for animals resistant to the AChR agonist levamisole. In conclusion, I will achieve a further understanding of the molecular mechanisms underlying ACh signaling at the NMJ by testing a novel role for FER-1/Dysferlin in the regulation of synaptic function and identifying novel genes required for regulation of post-synaptic ACh signaling. PUBLIC HEALTH RELEVANCE: Limb Girdle Muscular Dystrophy type 2B (LGMD2B), which is caused by loss of function mutations in the gene Dysferlin, results in progressive muscle weakness. Recent results suggest that loss of Dysferlin causes defects in synaptic function and this may contribute to LGMD2B phenotypes. Our research using C. elegans to study this novel pathophysiological mechanism of LGMD2B may lead to a better understanding of the molecular mechanisms that cause progressive muscle weakness and could lead to the development of new therapies to treat this incurable muscle disease.