Lysosomal Function of Progranulin and Neurodegeneration

项目摘要/摘要 原颗粒蛋白(PGRN)基因突变与两种不同的神经退行性疾病有关， 额颞叶变性(FTLD)和神经元蜡样脂褐素沉着症(NCL)。积累证据 提示PGRN在溶酶体中起着关键作用。然而，PGRN如何调节溶酶体功能和 防止神经退化的方法仍然难以捉摸。我们已经鉴定出丙皂苷(PSAP)，它是 作为PGRN结合伙伴的对溶酶体糖鞘糖脂降解必不可少的皂苷肽。我们 进一步表明，PGRN促进PSAP溶酶体通过神经元从细胞外间隙转运 贩卖受体山梨素。我们发现在PGRN缺陷小鼠中PSAP和皂苷的神经元水平降低 在因PGRN突变而导致的FTLD患者中。此外，我们还证明了PGRN与PSAP形成了络合物 还有溶酶体蛋白水解酶组织蛋白酶B和D，据报道组织蛋白酶D缺乏可导致FTLD。 相关病理和我们的初步研究表明，PSAP功能受损也会导致FTLD相关 小鼠的表型。因此，我们假设PGRN对于正确的PSAP和组织蛋白酶B和D是关键的 功能及PSAP和组织蛋白酶功能受损是FTLD-PGRN的一个重要发病机制。为了测试 在这一假设下，我们提出了三个具体目标。在Aim1中，我们将研究PGRN在调节 PSAP功能。将在WT和WT检测PSAP的贩运、加工和神经鞘糖脂代谢 PGRN-/-细胞和组织以及对照和FTLD-PGRN患者样本。在AIM2中，我们将确定 通过检测WT和PGRN-/-细胞和组织来研究PGRN在组织蛋白酶B和D转运和激活中的作用 以及对照和FTLD-PGRN患者样本。在Aim3中，我们将检测小鼠的FTLD样表型 用不同水平的PSAP或组织蛋白酶B或D来确定PSAP或组织蛋白酶的部分消失 功能可能有助于FTLD疾病的进展。此外，PSAP或组织蛋白酶B或D将是 通过腺相关病毒(AAV)过表达以确定PSAP或组织蛋白是否过表达 可以挽救与小鼠PGRN丢失相关的表型。总而言之，这些拟议的研究将摆脱 阐明PGRN如何调节溶酶体功能，并为研究糖尿病的发病机制提供新的见解 FTLD。我们希望我们的研究结果也能促进FTLD-PGRN的治疗发展 其他与PGRN有关的神经退行性疾病，如阿尔茨海默病。

PROJECT SUMMARY/ABSTRACT Mutations in the Progranulin (PGRN) gene have been linked to two distinct neurodegenerative diseases, frontotemporal lobar degeneration (FTLD) and neuronal ceroid lipofuscinosis (NCL). Accumulating evidence suggests a critical role of PGRN in the lysosome. However how PGRN regulates lysosomal function and protects against neurodegeneration remains elusive. We have identified prosaposin (PSAP), the precursor of saposin peptides essential for lysosomal glycosphingolipid degradation, as a PGRN binding partner. We further showed that PGRN facilitates PSAP lysosomal trafficking from the extracellular space via the neuronal trafficking receptor sortilin. We found reduced neuronal levels of PSAP and saposins in PGRN deficient mice and in FTLD patients due to PGRN mutations. Moreover, we showed that PGRN forms a complex with PSAP and lysosomal proteases cathepsin B and D. Cathepsin D deficiency has been reported to cause FTLD related pathology and our preliminary studies showed that impaired PSAP function also leads to FTLD related phenotypes in mice. Thus we hypothesize that PGRN is critical for proper PSAP and cathepsin B and D functions and impaired PSAP and cathepsin functions is one key disease mechanism of FTLD-PGRN. To test this hypothesis, we propose three specific aims. In Aim1, we will examine the role of PGRN in regulating PSAP function. PSAP trafficking, processing and glycosphingolipid metabolism will be assayed in WT and PGRN-/- cells and tissues as well as control and FTLD-PGRN patient samples. In Aim2, we will determine the role of PGRN in cathepsin B and D trafficking and activation by examining WT and PGRN-/- cells and tissues as well as control and FTLD-PGRN patient samples. In Aim3, we will assay FTLD like phenotypes in mice with different levels of PSAP or cathepsin B or D to determine whether partial loss of PSAP or cathepsin function could contribute to FTLD disease progression. Furthermore, PSAP or cathepsin B or D will be overexpressed via adeno associated viruses (AAV) to determine whether PSAP or cathepsin overexpression can rescue phenotypes associated with PGRN loss in mice. In summary, these proposed studies will shed light on how PGRN regulates lysosomal function and provide novel insights into the disease mechanism of FTLD. We expect the results from our studies to facilitate therapeutic development for FTLD-PGRN as well other neurodegenerative diseases with a reported role of PGRN, such as Alzheimer’s disease.