Background

Dysregulated ubiquitination modification occupies a pivotal role in hepatocellular carcinoma (HCC) tumorigenesis and progression. The ubiquitin aldehyde binding 1 (OTUB1) was aberrantly upregulated and exhibited the pro-tumorigenic function in HCC. However, the underlying mechanisms and responsible targets of OTUB1 remain unclear.

Methods

First, bioinformatics analysis, western blot and immunohistochemistry staining were applied to analyze OTUB1 expression in HCC specimens. Then, immunoprecipitation assay-tandem mass spectrometry (MS) combined with the gene set enrichment analysis (GSEA) was used to explore the downstream target of OTUB1. Co-immunoprecipitation and ubiquitination assays were used to identify the mechanisms involved. Finally, we explored the regulatory effect of MAZ on OTUB1 through ChIP-qPCR and dual-luciferase reporter assay.

Results

OTUB1 was broadly elevated in HCC tissues and promoted the proliferation and metastasis of HCC in vitro and in vivo. The receptor for activated C kinase 1 (RACK1) performed as a functional partner of OTUB1 and its hyperactivation was associated with aggressive development and other malignant features in HCC by activating oncogenes transcription. Mechanistically, OTUB1 directly bound to RACK1 at its C-terminal domain and decreased the K48-linked ubiquitination of RACK1 through its non-canonical suppression of ubiquitination activity, which stabilized RACK1 protein levels in HCC cells. Therefore, OTUB1 significantly increased multiple oncogenes expression and activated PI3K/AKT and FAK/ERK signaling in a RACK1-dependent manner in HCC. Moreover, the transcription factor MAZ upregulated OTUB1 expression through identifying a putative response element of OTUB1 promoter area.

Conclusions

Our findings might provide a new therapeutic strategy for HCC by modifying the MAZ-OTUB1-RACK1 axis.

中文翻译：

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OTUB1 通过其非典型泛素化稳定 RACK1，从而加速肝细胞癌的发生

背景

失调的泛素化修饰在肝细胞癌（HCC）肿瘤的发生和进展中起着关键作用。泛素醛结合 1 (OTUB1) 在 HCC 中异常上调并表现出促肿瘤功能。然而，OTUB1 的潜在机制和负责靶点仍不清楚。

方法

首先，应用生物信息学分析、蛋白质印迹和免疫组织化学染色分析HCC标本中OTUB1的表达。然后，采用免疫沉淀-串联质谱（MS）结合基因集富集分析（GSEA）来探索OTUB1的下游靶点。使用免疫共沉淀和泛素化测定来鉴定所涉及的机制。最后，我们通过 ChIP-qPCR 和双荧光素酶报告基因检测探讨了 MAZ 对 OTUB1 的调节作用。

结果

OTUB1在HCC组织中广泛升高，并在体外和体内促进HCC的增殖和转移。激活的 C 激酶 1 (RACK1) 受体作为 OTUB1 的功能伙伴，其过度激活通过激活癌基因转录与 HCC 的侵袭性发展和其他恶性特征相关。从机制上讲，OTUB1 直接与 RACK1 的 C 端结构域结合，并通过其对泛素化活性的非典型抑制来减少 RACK1 的 K48 连接泛素化，从而稳定 HCC 细胞中的 RACK1 蛋白水平。因此，在 HCC 中，OTUB1 显着增加多种癌基因的表达，并以 RACK1 依赖性方式激活 PI3K/AKT 和 FAK/ERK 信号传导。此外，转录因子 MAZ 通过识别 OTUB1 启动子区域的推定响应元件上调 OTUB1 表达。

结论

我们的研究结果可能通过修改 MAZ-OTUB1-RACK1 轴为 HCC 提供新的治疗策略。