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、溶酶体储存障碍(LSD)引起的疾病 由半乳糖脑苷酶(Galc)基因突变引起。Galc缺乏会导致脱髓鞘， 神经退化，并在几年内死亡。虽然存在延缓疾病的治疗方法，但也有 目前尚无治疗KD的有效方法。小鼠雪旺细胞特异性Galc(Galc-SC-CKO)缺失导致 KD的外周神经系统(PNS)病理表型，包括未消化的蓄积 底物伴有脱髓鞘、神经变性和一种非常类似的神经病 在KD患者中观察到。使用这个经过验证的Galc-SC-CKO模型，本研究将研究降解性 PNS中清除细胞底物的机制及其在发育和发病机制中的作用 Kd.初步结果表明，巨型自噬在疾病开始时增加，但随后停止。 并在疾病发展过程中下降。此外，自噬小体在雪旺细胞中的积累是 在电子显微镜(EM)下观察疾病进展。相比之下，由伴侣介导的 自噬途径似乎在KD的发病机制中没有显著贡献。我还发现溶酶体 转录因子TFEB在疾病进展过程中增加，表明串扰发生了变化 Galc-SC-CKO小鼠雪旺细胞的溶酶体与核之间的连接。因此，在这里，我建议研究 并在Galc-SC-CKO小鼠体内操纵巨噬细胞自噬和TFEB。有缺陷的自噬许可， 提示是自噬小体积累，伴随着TFEB调节的溶酶体的改变 生理可能导致底物堆积和溶酶体功能障碍，从而导致细胞 死于KD。了解降解途径中的特定机制变化有助于准确定位 KD和其他LSD中底物积累的原因和时间。这样的研究也可能与其他 更常见的疾病是由于溶酶体失灵、自噬和蛋白抑制。

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.