The hippo-YAP1/TEAD1-SLC7A5 axis: uncovering a novel therapeutic target for oxalate-induced renal tubular ferroptosis

To systematically investigate the regulatory mechanisms of ferroptosis in renal tubular epithelial cells under high oxalate stress, focusing on identifying key upstream signaling pathways and their therapeutic potential. We employed HK-2 cell cultures and Glyoxylate-induced mouse models of oxalate nephropathy. Multi-omics approaches including 4D-label-free proteomics, RNA-sequencing, and CUT&Tag were integrated to identify regulatory networks. Functional validation utilized genetic manipulation, pharmacological intervention, chromatin immunoprecipitation, and dual-luciferase reporter assays. High oxalate dose-dependently induced renal tubular ferroptosis and activated the Hippo pathway, leading to YAP1 phosphorylation and inactivation. Proteomic and multi-omics analyses identified YAP1 as a key regulator and SLC7A5 as its direct transcriptional target via TEAD1. High oxalate disrupted YAP1/TEAD1 binding to the SLC7A5 promoter, downregulating SLC7A5. Functional rescue confirmed that SLC7A5 loss mediated ferroptosis under YAP1 inhibition. Mechanistically, SLC7A5 downregulation restricted leucine availability and suppressed mTOR signaling, while leucine supplementation or mTOR reactivation reversed ferroptosis, demonstrating that SLC7A5 regulates ferroptosis via the leucine/mTOR axis. We establish the Hippo-YAP1/TEAD1-SLC7A5 axis as a master regulatory pathway controlling oxalate-induced ferroptosis. This pathway represents a promising therapeutic target for oxalate nephropathy and provides fundamental insights into stress-responsive ferroptosis regulation in kidney disease.

为系统探究高草酸胁迫下肾小管上皮细胞中铁死亡（ferroptosis）的调控机制，重点筛选关键上游信号通路并评估其治疗潜力。本研究采用HK-2细胞（HK-2 cell）培养体系与乙醛酸诱导的草酸肾病小鼠模型，整合4D无标记蛋白质组学（4D-label-free proteomics）、RNA测序（RNA-sequencing）及CUT&Tag技术（CUT&Tag）等多组学手段以解析调控网络。功能验证环节采用基因编辑、药物干预、染色质免疫沉淀（chromatin immunoprecipitation）及双荧光素酶报告基因实验（dual-luciferase reporter assays）等技术。研究发现，高草酸可剂量依赖性诱导肾小管上皮细胞发生铁死亡，并激活Hippo信号通路（Hippo pathway），致使YAP1蛋白（YAP1）发生磷酸化并失活。蛋白质组学及多组学分析显示，YAP1是核心调控因子，且通过TEAD1转录因子（TEAD1）将SLC7A5基因（SLC7A5）作为直接转录靶标。高草酸会破坏YAP1/TEAD1复合物与SLC7A5基因启动子的结合，从而下调SLC7A5的转录水平。功能挽救实验证实，在YAP1受抑制的情况下，SLC7A5的缺失会介导铁死亡的发生。机制研究表明，SLC7A5表达下调会限制亮氨酸（leucine）的可利用性并抑制mTOR信号通路（mTOR signaling），而补充亮氨酸或重新激活mTOR信号可逆转铁死亡现象，由此证明SLC7A5通过亮氨酸/mTOR信号轴调控铁死亡。本研究确立了Hippo-YAP1/TEAD1-SLC7A5信号轴作为调控草酸诱导铁死亡的核心通路，该通路可作为草酸肾病的潜在治疗靶点，同时为肾脏疾病中应激响应性铁死亡的调控机制提供了重要的理论基础。