Heat shot protein 90 (HSP90) AA1 functions as an onco-protein to regulate the assembly, manipulation, folding and degradation of its client proteins, including c-MYC. However, little is known about the mechanism of HSP90AA1 regulation. Transcriptome RNA-sequencing data of hepatocellular carcinoma (HCC) samples were used to detect the mRNA expression of FBXL6. Immunoprecipitation/Mass Spectrum (IP/MS) method was used to identify the interacting proteins of FBXL6. The co-immunoprecipitation assay was used to determine the interaction between FBXL6 and HSP90AA1. The in vivo ubiquitination assay was performed to determine the regulation of HSP90AA1 by FBXL6. Luciferase reporter and chromatin immunoprecipitation (ChIP) assays were used to determine the transcriptional regulation of FBXL6 by c-MYC. Immunohistochemical (IHC) staining was performed to study the correlation of FBXL6 and HSP90AA1 protein expression in 87 HCC samples. Cell counting and colony formation assays were implemented to detect the biological effects of FBXL6 on the growth of HCC cells in vitro. The effect of FBXL6 on HCC tumor growth in vivo was studied in a tumor xenograft model in mice. Here, we identified the orphan F-box protein FBXL6, a substrate recognition subunit of an SCF (Skp1-Cul1-F-box protein) complex, as the ubiquitin ligase for HSP90AA1. FBXL6 promoted K63-dependent ubiquitination of HSP90AA1 to stabilize it. Through analysis of the TCGA dataset, we found that FBXL6 was significantly increased in HCC tissues and positively correlated with c-MYC pathway. FBXL6 accumulation in HCC causes the stabilization and activation of c-MYC by preventing HSP90AA1 degradation. The activated c-MYC directly binds to the promoter region of FBXL6 to induce its mRNA expression. Collectively, our data revealed an unknown FBXL6-HSP90AA1-c-MYC axis which might contribute to the oncogenesis of HCC, and we propose that inhibition of FBXL6 might represent an effective therapeutic strategy for HCC treatment.

中文翻译：

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FBXL6 控制 c-MYC 通过泛素化和稳定 HSP90AA1 促进肝细胞癌。

热射蛋白 90 (HSP90) AA1 作为癌蛋白发挥作用，调节其客户蛋白（包括 c-MYC）的组装、操作、折叠和降解。然而，对 HSP90AA1 调节的机制知之甚少。肝细胞癌 (HCC) 样本的转录组 RNA 测序数据用于检测 FBXL6 的 mRNA 表达。免疫沉淀/质谱 (IP/MS) 方法用于鉴定 FBXL6 的相互作用蛋白。共免疫沉淀试验用于确定 FBXL6 和 HSP90AA1 之间的相互作用。进行体内泛素化测定以确定 FBXL6 对 HSP90AA1 的调节。荧光素酶报告基因和染色质免疫沉淀 (ChIP) 测定用于确定 c-MYC 对 FBXL6 的转录调控。进行免疫组织化学 (IHC) 染色以研究 87 个 HCC 样本中 FBXL6 和 HSP90AA1 蛋白表达的相关性。实施细胞计数和集落形成试验以检测 FBXL6 对体外 HCC 细胞生长的生物学影响。在小鼠的肿瘤异种移植模型中研究了 FBXL6 对体内 HCC 肿瘤生长的影响。在这里，我们鉴定了孤儿 F-box 蛋白 FBXL6，一种 SCF（Skp1-Cul1-F-box 蛋白）复合物的底物识别亚基，作为 HSP90AA1 的泛素连接酶。FBXL6 促进 HSP90AA1 的 K63 依赖性泛素化以稳定它。通过对TCGA数据集的分析，我们发现FBXL6在HCC组织中显着增加，并且与c-MYC通路呈正相关。FBXL6 在 HCC 中的积累通过阻止 HSP90AA1 降解导致 c-MYC 的稳定和激活。活化的 c-MYC 直接与 FBXL6 的启动子区域结合以诱导其 mRNA 表达。总的来说，我们的数据揭示了一个未知的 FBXL6-HSP90AA1-c-MYC 轴，它可能有助于 HCC 的发生，我们提出抑制 FBXL6 可能代表 HCC 治疗的有效治疗策略。