How lysosome and MTORC1 signaling interact remains elusive in terminally differentiated cells. A G4C2 repeat expansion in C9orf72 is the most common cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) (C9ALS-FTD). We previously identified a C9orf72-SMCR8-containing complex. Here we found that c9orf72 and smcr8 double-knockout (dKO) mice exhibit similar but more severe immune defects than the individual knockouts. In c9orf72 or smcr8 mutant macrophages, lysosomal degradation and exocytosis were impaired due to the disruption of autolysosome acidification. As a result of impaired lysosomal degradation, MTOR protein was aberrantly increased, resulting in MTORC1 signaling overactivation. Inhibition of hyperactive MTORC1 partially rescued macrophage dysfunction, splenomegaly and lymphadenopathy in c9orf72 or smcr8 mutant mice. Pharmacological inhibition of lysosomal degradation upregulated MTOR protein and MTORC1 signaling in differentiated wild-type macrophages, which resemble phenotypes in KO mice. In contrast, C9orf72 or Smcr8 depletion in proliferating macrophages decreased MTORC1 signaling. Our studies causatively link C9orf72-SMCR8’s cellular functions in lysosomal degradation, exocytosis, and MTORC1 signaling with their organism-level immune regulation, suggesting cell state (proliferation vs. differentiation)-dependent regulation of MTOR signaling via lysosomes.Abbreviations: ALS: amyotrophic lateral sclerosis; ATG13: autophagy related 13; BMDMs: bone marrow-derived macrophages; BafA₁: bafilomycin A₁; C9orf72: C9orf72, member of C9orf72-SMCR8 complex; CD68: CD68 antigen; ConA: concanamycin A; dKO: double knockout; DENN: differentially expressed in normal and neoplastic cells; FTD: frontotemporal dementia; GEF: guanine nucleotide exchange factor; IFNB1: interferon beta 1, fibroblast; IFNG: interferon gamma; IL1B/IL-1β: interleukin 1 beta; IL6: interleukin 6; iPSCs: induced pluripotent stem cells; LAMP1: lysosomal-associated membrane protein 1; LPOs: LAMP1-positive organelles; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; LPS: lipopolysaccharide; MTORC1: mechanistic target of rapamycin kinase complex 1; MEFs: mouse embryonic fibroblasts; MNs: motor neurons; NOS2/iNOS: nitric oxide synthase 2, inducible; RAN: repeat-associated non-AUG; RB1CC1/FIP200: RB1-inducible coiled-coil 1; RPS6/S6: ribosomal protein S6; RPS6KB1/S6K1: ribosomal protein S6 kinase, polypeptide 1; SMCR8: Smith-Magenis syndrome chromosome region, candidate 8; SQSTM1/p62: sequestosome 1; TFEB: transcription factor EB; TNF: tumor necrosis factor; TSC1: TSC complex subunit 1; ULK1: unc-51 like kinase 1; v-ATPase: vacuolar-type H⁺-translocating ATPase

在终末分化的细胞中，溶酶体和 MTORC1 信号如何相互作用仍然难以捉摸。在甲G4C2重复扩张C9orf72是最常见的原因家族性肌萎缩侧索硬化症（ALS）和额颞叶痴呆（FTD）（C9ALS-FTD）。我们之前确定了一个包含 C9orf72-SMCR8 的复合物。在这里，我们发现c9orf72和smcr8双基因敲除 (dKO) 小鼠表现出与单个基因敲除相似但更严重的免疫缺陷。在c9orf72或smcr8由于自溶酶体酸化的破坏，突变巨噬细胞、溶酶体降解和胞吐作用受损。由于溶酶体降解受损，MTOR 蛋白异常增加，导致 MTORC1 信号过度激活。抑制过度活跃的 MTORC1 部分挽救了c9orf72或smcr8突变小鼠的巨噬细胞功能障碍、脾肿大和淋巴结病。溶酶体降解的药理学抑制上调了分化的野生型巨噬细胞中的 MTOR 蛋白和 MTORC1 信号，这类似于 KO 小鼠的表型。相比之下，C9orf72或Smcr8增殖巨噬细胞的耗竭降低了 MTORC1 信号传导。我们的研究将 C9orf72-SMCR8 在溶酶体降解、胞吐作用和 MTORC1 信号传导中的细胞功能与其生物体水平的免疫调节相关联，表明细胞状态（增殖与分化）依赖于通过溶酶体调节 MTOR 信号传导。缩写：ALS：肌萎缩侧索硬化；ATG13：自噬相关13；BMDMs：骨髓来源的巨噬细胞；BafA ₁ : 巴弗洛霉素 A ₁; C9orf72：C9orf72，C9orf72-SMCR8复合体的成员；CD68：CD68抗原；ConA：刀豆霉素A；dKO：双淘汰赛；DENN：在正常和肿瘤细胞中差异表达；FTD：额颞叶​​痴呆；GEF：鸟嘌呤核苷酸交换因子；IFNB1：干扰素β1，成纤维细胞；IFNG：干扰素γ；IL1B/IL-1β：白细胞介素 1β；IL6：白介素 6；iPSCs：诱导多能干细胞；LAMP1：溶酶体相关膜蛋白 1；LPO：LAMP1 阳性细胞器；MAP1LC3/LC3：微管相关蛋白1轻链3；LPS：脂多糖；MTORC1：雷帕霉素激酶复合物 1 的机制靶点；MEFs：小鼠胚胎成纤维细胞；MNs：运动神经元；NOS2/iNOS：一氧化氮合酶 2，诱导型；RAN：重复相关的非AUG；RB1CC1/FIP200：RB1-可感应盘绕线圈1；RPS6/S6：核糖体蛋白S6；RPS6KB1/S6K1：核糖体蛋白 S6 激酶，多肽 1；SMCR8：Smith-Magenis 综合征染色体区域，候选 8；SQSTM1/p62：sequestosome 1；TFEB：转录因子EB；TNF：肿瘤坏死因子；TSC1：TSC复合亚基1；ULK1：unc-51 像激酶 1；v-ATPase：液泡型 H⁺-易位 ATP 酶