STK11/LKB1突变型NSCLC中KEAP1的共存突变激活NFE2L2/NRF2以补偿代谢适应期间STK11-AMPK活性的丧失。表征代谢应激期间STK11-AMPK和KEAP1-NFE2L2途径之间的调节串扰对于理解共同发生的突变的含义至关重要。这里,我们发现代谢应激增加了SQSTM1/p62的表达和磷酸化,这对于NFE2L2和AMPK的激活至关重要,协同抗氧化防御和肿瘤生长。SQSTM1驱动的NFE2L2和AMPK的双重激活是通过诱导KEAP1的宏观自噬/自噬降解并促进溶酶体膜上的AXIN-STK11-AMPK复合物形成来实现的,分别。相比之下,代谢应激诱导的SQSTM1表达和磷酸化也需要STK11-AMPK活性,提示AMPK和SQSTM1之间存在双正反馈环.机械上,TFEB和TFE3的PPP2/PP2A依赖性去磷酸化增加了SQSTM1的表达,这是由低糖代谢和AMPK依赖性质子还原引起的溶酶体脱酸诱导的。此外,SQSTM1磷酸化被MAP3K7/TAK1增加,其被ROS和pH依赖性分泌的溶酶体Ca2+激活。重要的是,SQSTM1在S24和S226的磷酸化对于AMPK和NFE2L2的激活至关重要。值得注意的是,乳酸提供的质子消除了代谢应激引起的影响。总的来说,我们的数据揭示了AMPK和SQSTM1之间的一个新的双正反馈回路,导致AMPK和NFE2L2的双重激活,这可能解释了STK11和KEAP1发生共同突变的原因,并为肺癌提供了有希望的治疗策略.
Co-occurring mutations in
KEAP1 in STK11/LKB1-mutant NSCLC activate NFE2L2/NRF2 to compensate for the loss of STK11-AMPK activity during metabolic adaptation. Characterizing the regulatory crosstalk between the STK11-AMPK and
KEAP1-NFE2L2 pathways during metabolic stress is crucial for understanding the implications of co-occurring mutations. Here, we found that metabolic stress increased the expression and phosphorylation of SQSTM1/p62, which is essential for the activation of NFE2L2 and AMPK, synergizing antioxidant defense and tumor growth. The SQSTM1-driven dual activation of NFE2L2 and AMPK was achieved by inducing macroautophagic/autophagic degradation of
KEAP1 and facilitating the AXIN-STK11-AMPK complex formation on the lysosomal membrane, respectively. In contrast, the STK11-AMPK activity was also required for metabolic stress-induced expression and phosphorylation of SQSTM1, suggesting a double-positive feedback loop between AMPK and SQSTM1. Mechanistically, SQSTM1 expression was increased by the PPP2/PP2A-dependent dephosphorylation of TFEB and TFE3, which was induced by the lysosomal deacidification caused by low glucose metabolism and AMPK-dependent proton reduction. Furthermore, SQSTM1 phosphorylation was increased by MAP3K7/TAK1, which was activated by ROS and pH-dependent secretion of lysosomal Ca2+. Importantly, phosphorylation of SQSTM1 at S24 and S226 was critical for the activation of AMPK and NFE2L2. Notably, the effects caused by metabolic stress were abrogated by the protons provided by lactic acid. Collectively, our data reveal a novel double-positive feedback loop between AMPK and SQSTM1 leading to the dual activation of AMPK and NFE2L2, potentially explaining why co-occurring mutations in STK11 and
KEAP1 happen and providing promising therapeutic strategies for lung cancer.Abbreviations: AMPK: AMP-activated protein kinase; BAF1: bafilomycin A1; ConA: concanamycin A; DOX: doxycycline; IP: immunoprecipitation;
KEAP1: kelch like ECH associated protein 1; LN: low nutrient; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MCOLN1/TRPML1: mucolipin TRP cation channel 1; MEFs: mouse embryonic fibroblasts; MTORC1: mechanistic target of rapamycin kinase complex 1; NAC: N-acetylcysteine; NFE2L2/NRF2: NFE2 like bZIP transcription factor 2; NSCLC: non-small cell lung cancer; PRKAA/AMPKα: protein kinase AMP-activated catalytic subunit alpha; PPP2/PP2A: protein phosphatase 2; ROS: reactive oxygen species; PPP3/calcineurin: protein phosphatase 3; RPS6KB1/p70S6K: ribosomal protein S6 kinase B1; SQSTM1/p62: sequestosome 1; STK11/LKB1: serine/threonine kinase 11; TCL: total cell lysate; TFEB: transcription factor EB; TFE3: transcription factor binding to IGHM enhancer 3; V-ATPase: vacuolar-type H+-translocating ATPase.