Translational Research Center for Protein Function Control, Yonsei University, Seoul, Korea.
Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University, Seoul, Korea.
Cell Commun Signal. 2019 Jul 30;17(1):88. doi: 10.1186/s12964-019-0403-x.
Stabilization of RAS is a key event for the hyper-activation of Wnt/β-catenin signaling and activation of cancer stem cell (CSC) in colorectal cancer (CRC). WD Repeat protein 76 (WDR76) mediates the polyubiquitination-dependent degradation of RAS in hepatocellular carcinoma (HCC). We investigated whether WDR76 destabilizes RAS and acts as a tumor suppressor inhibiting CSC activation in CRC.
We generated mice with deletion of Wdr76 (Wdr76) and crosses of Wdr76 with Apc (Wdr76; Apc) and compared them with wildtype mice (Wdr76) and Apc mice (Wdr76; Apc), respectively. Intestinal crypt lengthening, tumorigenesis and CSC activation were analyzed by histology, immunohistochemistry, and immunoblotting. CRC cell line was engineered to stably express or knockdown WDR76 or control vector and was analyzed after spheroid culture.
Wdr76 mice, with increased Ras level, displayed crypt elongation and hyper-proliferation. Wdr76; Apc mice developed more tumors with bigger sizes than Apc mice and their tumors showed increased proliferation and CSC activation with elevated RAS and β-catenin levels. In CRC cells, overexpression or knockdown of WDR76 decreased or increased the numbers and sizes of CRC spheroids with inhibition or activation of CSC markers, respectively. In human CRC, lower level of WDR76 was associated with poor patient survival.
In analyses of mice with deletion of Wdr76 and CRC spheroids, we found that RAS stability plays important roles in tumorigenesis by affecting proliferation and CSC activation. Our results suggest that destabilization of RAS by WDR76 is a potential strategy for targeting malignant CRC involving CSC activation.
RAS 的稳定是结直肠癌（CRC）中 Wnt/β-连环蛋白信号的过度激活和癌症干细胞（CSC）激活的关键事件。WD 重复蛋白 76（WDR76）介导了肝癌（HCC）中 RAS 的多泛素化依赖性降解。我们研究了 WDR76 是否使 RAS 不稳定并作为肿瘤抑制因子抑制 CRC 中的 CSC 激活。
我们生成了 Wdr76 缺失（Wdr76）的小鼠，并进行了 Wdr76 与 Apc（Wdr76；Apc）的杂交，分别与野生型小鼠（Wdr76）和 Apc 小鼠（Wdr76；Apc）进行比较。通过组织学、免疫组织化学和免疫印迹分析肠隐窝伸长、肿瘤发生和 CSC 激活。对 CRC 细胞系进行了稳定表达或敲低 WDR76 或对照载体的工程改造，并在球体培养后进行了分析。
Wdr76 小鼠，Ras 水平增加，表现出隐窝伸长和过度增殖。Wdr76；Apc 小鼠比 Apc 小鼠形成更多更大的肿瘤，其肿瘤显示出更高的增殖和 CSC 激活，伴有 RAS 和 β-连环蛋白水平的升高。在 CRC 细胞中，过表达或敲低 WDR76 分别减少或增加了 CRC 球体的数量和大小，抑制或激活了 CSC 标志物。在人 CRC 中，WDR76 水平较低与患者预后不良相关。
在 Wdr76 缺失的小鼠和 CRC 球体分析中，我们发现 RAS 稳定性通过影响增殖和 CSC 激活在肿瘤发生中起重要作用。我们的结果表明，WDR76 使 RAS 不稳定可能是一种针对涉及 CSC 激活的恶性 CRC 的潜在策略。
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