Analysis of the tumor suppressor gene Tsc1 in motor neuron patterning

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): Motor neurons (MNs) enable control over locomotion, respiration and autonomic responses, and are profoundly affected by diseases such as spinal muscle atrophy (SMA) and amyotrophic lateral sclerosis (ALS). In vertebrates, MNs develop in the ventral spinal cord and hindbrain, their cell bodies migrate to specific positions along the medio-lateral and dorso-ventral axes of the CNS and their axons exit the CNS to established precise and stereotypic innervation of muscle targets. Although numerous guidance molecules have been identified that play an essential role in MN axon navigation, our mechanistic understanding of the signaling pathways that control MN axon steering at specific choice points remains fragmentary. The long-term goal of the proposed work is to identify novel mechanisms that control MN axon navigation in vivo. As an unbiased approach to identify such genes, a forward genetic screen was carried out using a reporter mouse with GFP-labeled MN axons and td-tomato-labeled MN nuclei. From the ENU mutagenesis screen, nine independent mutants were identified and their corresponding genes were cloned. Interestingly, several of these mutants display axon targeting defects in which ventral-projecting MN axons aberrantly project dorsally into the sensory ganglia. The MN pathfinding defects in a mouse mutant called Greenlight (GrL) are caused by a missense mutation in the tuberous sclerosis complex 1 (TSC1) gene. The objective in this application is to understand how the identified GrL/TSC1 mutation contributes to MN pathfinding defects. Mutations in human TSC1 cause a spectrum of neurological phenotypes including benign tumors, epilepsy and autism; however the pathophysiology of these symptoms is poorly understood. The central hypothesis is that TSC1, in conjunction with TSC2, negatively regulates the mTOR pathway downstream of ephrin/Eph in developing MNs. This hypothesis has been developed based on the identification of the GrL/TSC1 mutation in my screen, and the recent observation that inhibitory axon guidance cues negatively regulate the mTOR pathway. The rationale for the proposed research is that a detailed understanding of TSC1 function has the potential to translate into improved therapeutic interventions in patients suffering from tuberous sclerosis. Guided by strong preliminary data, the hypothesis will be tested by pursuing two specific aims: 1) to study how the identified mutation in GrL/TSC1 influences TSC1/TSC2 complex formation and to analyze in detail how the GrL/TSC1 mutation affects MN and sensory axon patterning in the developing PNS. 2) to study whether ephrin/EphA operates upstream of TSC1 and whether altered regulation of the TOR complex 1 downstream of TSC1 results in the GrL MN pathfinding defects. The proposed experiments are innovative because they are based on the identification of the GrL/TSC1 allele as a novel mutation that causes MN pathfinding defects. Ultimately, the knowledge gained has the potential to inform how the tumor suppressor gene TSC1 functions in nervous system development and how perturbation of this pathway can cause the neurological manifestations typically observed in TSC patients.