Understanding the Role of Autophagy and Transcription Factor EB (TFEB) in Krabbe Disease Pathogenesis

项目摘要 Krabbe病（KD）是一种遗传性常染色体隐性遗传、衰弱性、溶酶体贮积症（LSD）， 半乳糖苷酶（Galc）基因突变。Galc缺乏导致脱髓鞘， 神经退化，几年内死亡虽然存在延迟疾病的治疗方法， 目前还没有治疗KD的方法。雪旺细胞特异性缺失小鼠Galc（Galc-SC-cKO）导致 KD的外周神经系统（PNS）病理表型，包括未消化的 伴有脱髓鞘、神经变性和与之非常相似的神经病变的基质 在KD患者中观察到。使用该经验证的Galc-SC-cKO模型，本研究将研究 清除PNS中细胞底物的机制，以及它们如何促进发展和发病机制 KD的。初步结果表明，巨自噬在疾病开始时增加， 并在疾病进展期间降低。此外，自噬体在雪旺细胞中的积累是 疾病进展期间在电子显微镜（EM）下观察。相反，伴侣介导的 自噬途径似乎对KD发病机制没有显著贡献。我还发现溶酶体 一种转录因子TFEB在疾病进展过程中增加，表明串扰的改变， 在来自Galc-SC-cKO小鼠的许旺细胞中，溶酶体与细胞核之间的差异。因此，在这里，我建议研究 并在体内操纵Galc-SC-cKO小鼠中的巨自噬和TFEB。自噬清除缺陷， 提示自噬体的积累，伴随着改变的TFEB调节的溶酶体 生理学可能导致底物积累和溶酶体功能障碍，导致细胞凋亡。 KD的死亡了解降解途径中的具体机制变化可以帮助确定 KD和其他LSD中底物蓄积的原因和时间。这些研究也可能与其他研究有关。 更常见的疾病是由溶酶体、自噬和蛋白质稳定的失败引起的。

Project Summary Krabbe disease (KD) is an inherited autosomal recessive, debilitating, lysosomal storage disorder (LSD) caused by mutations in galactosylcerebrosidase (Galc) gene. Galc deficiency leads to demyelination, neurodegeneration, and death within a couple years. Although therapies that delay the disease exist, there is currently no available cure for KD. Schwann cells-specific deletion of Galc (Galc-SC-cKO) in mice leads to a Peripheral Nervous System (PNS) pathological phenotype of KD, which includes accumulation of undigested substrates accompanied by demyelination, neurodegeneration, and a neuropathy very similar to the one observed in KD patients. Using this validated Galc-SC-cKO model, this study will investigate the degradative mechanisms that clear cellular substrate in the PNS, and how they contribute to development and pathogenesis of KD. Preliminary results indicate that macroautophagy is increased at the beginning of the disease, but it halts and decreases during disease progression. In addition, autophagosome accumulation in Schwann cells is observed under electron microscopy (EM) during disease progression. In contrast, chaperone-mediated autophagy pathway does not appear to contribute significantly to KD pathogenesis. I also find that the lysosomal transcription factor, TFEB, is increased during disease progression, suggesting an alteration in crosstalk between the lysosome and the nucleus in Schwann cells from Galc-SC-cKO mice. Thus, here I propose to study and manipulate macroautophagy and TFEB in Galc-SC-cKO mice in vivo. Defective autophagic clearance, suggested by accumulation of autophagosomes, accompanied by an altered TFEB-regulated lysosomal physiology may be leading to substrate accumulation and to lysosomal dysfunction, contributing to cellular demise in KD. Understanding specific mechanistic changes in degradative pathways could help pinpoint the cause and timing of substrate accumulation in KD and other LSDs. Such studies may also be relevant to other more common diseases caused by failure in lysosomes, autophagy, and proteostasis.