Abstract：
BACKGROUND: Deregulation of cell death is a common characteristic of cancer, and resistance to this process often occurs in lung cancer. Understanding the molecular mechanisms underlying an aberrant cell death is important. Recent studies have emphasized the involvement of calmodulin-regulated spectrin-associated protein 3 (CAMSAP3) in lung cancer aggressiveness, its influence on cell death regulation remains largely unexplored.
METHODS: CAMSAP3 was knockout in lung cancer cells using CRISPR-Cas9 system. Cell death and autophagy were evaluated using MTT and autophagic detection assays. Protein interactions were performed by proteomic analysis and immunoprecipitation. Protein expressions and their cytoplasmic localization were analyzed through immunoblotting and immunofluorescence techniques.
RESULTS: This study reveals a significant correlation between low CAMSAP3 expression and poor overall survival rates in lung cancer patients. Proteomic analysis identified high mobility group box 1 (HMGB1) as a candidate interacting protein involved in the regulation of cell death. Treatment with trichostatin A (TSA), an inhibitor of histone deacetylases (HDACs) resulted in increased HMGB1 acetylation and its translocation to the cytoplasm and secretion, thereby inducing autophagic cell death. However, this process was diminished in CAMSAP3 knockout lung cancer cells. Mechanistically, immunoprecipitation indicated an interaction between CAMSAP3 and HMGB1, particularly with its acetylated form, in which this complex was elevated in the presence of TSA.
CONCLUSIONS: CAMSAP3 is prerequisite for TSA-mediated autophagic cell death by interacting with cytoplasmic acetylated HMGB1 and enhancing its release.
CONCLUSIONS: This finding provides molecular insights into the role of CAMSAP3 in regulating cell death, highlighting its potential as a therapeutic target for lung cancer treatment.
METHODS: CAMSAP3 was knockout in lung cancer cells using CRISPR-Cas9 system. Cell death and autophagy were evaluated using MTT and autophagic detection assays. Protein interactions were performed by proteomic analysis and immunoprecipitation. Protein expressions and their cytoplasmic localization were analyzed through immunoblotting and immunofluorescence techniques.
RESULTS: This study reveals a significant correlation between low CAMSAP3 expression and poor overall survival rates in lung cancer patients. Proteomic analysis identified high mobility group box 1 (HMGB1) as a candidate interacting protein involved in the regulation of cell death. Treatment with trichostatin A (TSA), an inhibitor of histone deacetylases (HDACs) resulted in increased HMGB1 acetylation and its translocation to the cytoplasm and secretion, thereby inducing autophagic cell death. However, this process was diminished in CAMSAP3 knockout lung cancer cells. Mechanistically, immunoprecipitation indicated an interaction between CAMSAP3 and HMGB1, particularly with its acetylated form, in which this complex was elevated in the presence of TSA.
CONCLUSIONS: CAMSAP3 is prerequisite for TSA-mediated autophagic cell death by interacting with cytoplasmic acetylated HMGB1 and enhancing its release.
CONCLUSIONS: This finding provides molecular insights into the role of CAMSAP3 in regulating cell death, highlighting its potential as a therapeutic target for lung cancer treatment.
摘要：
背景：细胞死亡失调是癌症的共同特征，对这一过程的抵抗通常发生在肺癌中。了解异常细胞死亡的分子机制很重要。最近的研究强调钙调蛋白调节的血影蛋白相关蛋白3（CAMSAP3）参与肺癌的侵袭性，其对细胞死亡调节的影响在很大程度上仍未被探索。
方法：使用CRISPR-Cas9系统在肺癌细胞中敲除CAMSAP3。使用MTT和自噬检测测定法评估细胞死亡和自噬。通过蛋白质组学分析和免疫沉淀进行蛋白质相互作用。通过免疫印迹和免疫荧光技术分析蛋白质表达及其细胞质定位。
结果：本研究揭示了肺癌患者低CAMSAP3表达与低总生存率之间的显著相关性。蛋白质组学分析鉴定了高迁移率基团盒1（HMGB1）作为参与细胞死亡调节的候选相互作用蛋白。曲古霉素A（TSA）治疗，组蛋白去乙酰化酶（HDACs）的抑制剂导致HMGB1乙酰化增加，并易位到细胞质和分泌，从而诱导自噬性细胞死亡。然而,在CAMSAP3敲除的肺癌细胞中,这一过程减弱.机械上,免疫沉淀表明CAMSAP3和HMGB1之间的相互作用，特别是其乙酰化形式，其中该复合物在TSA存在下升高。
结论：CAMSAP3通过与细胞质乙酰化HMGB1相互作用并增强其释放，是TSA介导的自噬性细胞死亡的先决条件。
结论：这一发现为CAMSAP3在调节细胞死亡中的作用提供了分子见解，强调其作为肺癌治疗靶点的潜力。
方法：使用CRISPR-Cas9系统在肺癌细胞中敲除CAMSAP3。使用MTT和自噬检测测定法评估细胞死亡和自噬。通过蛋白质组学分析和免疫沉淀进行蛋白质相互作用。通过免疫印迹和免疫荧光技术分析蛋白质表达及其细胞质定位。
结果：本研究揭示了肺癌患者低CAMSAP3表达与低总生存率之间的显著相关性。蛋白质组学分析鉴定了高迁移率基团盒1（HMGB1）作为参与细胞死亡调节的候选相互作用蛋白。曲古霉素A（TSA）治疗，组蛋白去乙酰化酶（HDACs）的抑制剂导致HMGB1乙酰化增加，并易位到细胞质和分泌，从而诱导自噬性细胞死亡。然而,在CAMSAP3敲除的肺癌细胞中,这一过程减弱.机械上,免疫沉淀表明CAMSAP3和HMGB1之间的相互作用，特别是其乙酰化形式，其中该复合物在TSA存在下升高。
结论：CAMSAP3通过与细胞质乙酰化HMGB1相互作用并增强其释放，是TSA介导的自噬性细胞死亡的先决条件。
结论：这一发现为CAMSAP3在调节细胞死亡中的作用提供了分子见解，强调其作为肺癌治疗靶点的潜力。
