The Affiliated Hospital of Guilin Medical University, Guangxi Key Laboratory of Brain and Cognitive Neuroscience, Guangxi Neurological Diseases Clinical Research Center, Guilin, Guangxi, China.
Department of Immunobiology, Yale University School of Medicine, New Haven, CT, USA.
Autophagy. 2021 Mar;17(3):723-742. doi: 10.1080/15548627.2020.1731266. Epub 2020 Mar 18.
Although the treatment of brain tumors by targeting kinase-regulated macroautophagy/autophagy, is under investigation, the precise mechanism underlying autophagy initiation and its significance in glioblastoma (GBM) remains to be defined. Here, we report that PAK1 (p21 [RAC1] activated kinase 1) is significantly upregulated and promotes GBM development. The Cancer Genome Atlas analysis suggests that the oncogenic role of PAK1 in GBM is mainly associated with autophagy. Subsequent experiments demonstrate that PAK1 indeed serves as a positive modulator for hypoxia-induced autophagy in GBM. Mechanistically, hypoxia induces ELP3-mediated PAK1 acetylation at K420, which suppresses the dimerization of PAK1 and enhances its activity, thereby leading to subsequent PAK1-mediated ATG5 (autophagy related 5) phosphorylation at the T101 residue. This event not only protects ATG5 from ubiquitination-dependent degradation but also increases the affinity between the ATG12-ATG5 complex and ATG16L1 (autophagy related 16 like 1). Consequently, ELP3-dependent PAK1 (K420) acetylation and PAK1-mediated ATG5 (T101) phosphorylation are required for hypoxia-induced autophagy and brain tumorigenesis by promoting autophagosome formation. Silencing with shRNA or small molecule inhibitor FRAX597 potentially blocks autophagy and GBM growth. Furthermore, SIRT1-mediated PAK1-deacetylation at K420 hinders autophagy and GBM growth. Clinically, the levels of PAK1 (K420) acetylation significantly correlate with the expression of ATG5 (T101) phosphorylation in GBM patients. Together, this report uncovers that the acetylation modification and kinase activity of PAK1 plays an instrumental role in hypoxia-induced autophagy initiation and maintaining GBM growth. Therefore, PAK1 and its regulator in the autophagy pathway might represent potential therapeutic targets for GBM treatment. 3-MA: 3-methyladenine; Ac-CoA: acetyl coenzyme A; ATG5: autophagy related 5; ATG16L1, autophagy related 16 like 1; BafA: bafilomycin A; CDC42: cell division cycle 42; CGGA: Chinese Glioma Genome Atlas; CHX, cycloheximide; ELP3: elongator acetyltransferase complex subunit 3; GBM, glioblastoma; HBSS: Hanks balanced salts solution; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MAP2K1: mitogen-activated protein kinase kinase 1; MAPK14, mitogen-activated protein kinase 14; PAK1: p21 (RAC1) activated kinase 1; PDK1: pyruvate dehydrogenase kinase 1; PGK1, phosphoglycerate kinase 1; PTMs: post-translational modifications; RAC1: Rac family small GTPase 1; SQSTM1: sequestosome 1; TCGA, The Cancer Genome Atlas.
虽然通过靶向激酶调节的宏观自噬/自噬来治疗脑肿瘤正在研究中，但自噬起始的精确机制及其在胶质母细胞瘤 (GBM) 中的意义仍有待确定。在这里，我们报告 PAK1（p21[RAC1]激活激酶 1）显着上调并促进 GBM 发展。癌症基因组图谱分析表明，PAK1 在 GBM 中的致癌作用主要与自噬有关。随后的实验表明，PAK1 确实是 GBM 中缺氧诱导自噬的正调节剂。在机制上，缺氧诱导 ELP3 介导的 PAK1 在 K420 处乙酰化，抑制 PAK1 二聚化并增强其活性，从而导致随后 PAK1 介导的 ATG5（自噬相关 5）在 T101 残基处磷酸化。该事件不仅保护 ATG5 免受泛素依赖性降解，而且增加 ATG12-ATG5 复合物与 ATG16L1（自噬相关 16 样 1）之间的亲和力。因此，ELP3 依赖性 PAK1（K420）乙酰化和 PAK1 介导的 ATG5（T101）磷酸化对于通过促进自噬体形成来诱导缺氧诱导的自噬和脑肿瘤发生是必需的。用 shRNA 或小分子抑制剂 FRAX597 沉默可能会阻止自噬和 GBM 生长。此外，SIRT1 介导的 PAK1 在 K420 处去乙酰化会阻碍自噬和 GBM 生长。临床上，PAK1（K420）乙酰化水平与 GBM 患者中 ATG5（T101）磷酸化的表达显着相关。总的来说，本报告揭示了 PAK1 的乙酰化修饰和激酶活性在缺氧诱导的自噬起始和维持 GBM 生长中起着重要作用。因此，PAK1 及其在自噬途径中的调节剂可能代表治疗 GBM 的潜在治疗靶标。3-MA：3-甲基腺嘌呤；乙酰辅酶 A：乙酰辅酶 A；ATG5：自噬相关 5；ATG16L1，自噬相关 16 样 1；BafA：巴佛霉素 A；CDC42：细胞分裂周期 42；CGGA：中国胶质瘤基因组图谱；CHX，环己酰亚胺；ELP3：延伸乙酰转移酶复合物亚基 3；GBM，胶质母细胞瘤；HBSS：汉克斯平衡盐溶液；MAP1LC3B/LC3：微管相关蛋白 1 轻链 3β；MAP2K1：丝裂原活化蛋白激酶激酶 1；MAPK14，丝裂原活化蛋白激酶 14；PAK1：p21（RAC1）激活激酶 1；PDK1：丙酮酸脱氢酶激酶 1；PGK1，磷酸甘油酸激酶 1；PTMs：翻译后修饰；RAC1：Rac 家族小 GTP 酶 1；SQSTM1：自噬体 1；TCGA，癌症基因组图谱。
