KCTD9 inhibits the Wnt/β-catenin pathway by decreasing the level of β-catenin in colorectal cancer

Colorectal cancer (CRC) is the second leading cause of cancer mortality worldwide. However, the molecular mechanisms underlying CRC progression remain to be further defined to improve patient outcomes. In this study, we found that KCTD9, a member of the potassium channel tetramerization domain-containing (KCTD) gene family, was commonly downregulated in CRC tissues and that KCTD9 expression was negatively correlated with the clinical CRC stage. Survival analysis showed that patients whose tumors expressed low KCTD9 levels had poorer outcomes. Functional analyses revealed that KCTD9 overexpression inhibited CRC cell proliferation and metastasis, whereas KCTD9 knockdown promoted CRC cell proliferation and metastasis in both in vitro and in vivo models. Manipulating KCTD9 levels in CRC cells via overexpression or knockdown showed KCTD9 expression positively influenced the degradation of β-catenin levels leading to inhibition of Wnt signaling and reductions in Wnt pathway target gene expression. Mechanistically, we found KCTD9 associated with ZNT9 (Zinc Transporter 9), a coactivator of β-catenin-mediated gene transcription. The overexpression of KCTD9 or knockdown of ZNT9 in CRC cells increased the polyubiquitination and proteasomal degradation of β-catenin. In turn, the KCTD9-ZNT9 interaction disrupted interactions between β-catenin and ZNT9, thereby leading to decreased β-catenin target gene expression and the inhibition of Wnt signaling. In conclusion, our findings propose that KCTD9 functions as a tumor suppressor that inhibits CRC cell proliferation and metastasis by inactivating the Wnt/β-catenin pathway. Moreover, its frequent downregulation in CRC suggests KCTD9 as a potential prognostic and therapeutic target in CRC. Comparative gene expression profiling analysis of RNA-seq data for adjacent normal colonic mucosa, primary colon adenocarcinoma and distant liver metastasis

结直肠癌（Colorectal cancer, CRC）是全球范围内排名第二的癌症致死病因。然而，结直肠癌进展的潜在分子机制仍有待进一步阐明，以改善患者的临床结局。本研究发现，钾通道四聚化结构域包含（potassium channel tetramerization domain-containing, KCTD）基因家族成员KCTD9在结直肠癌组织中普遍下调，且KCTD9的表达水平与结直肠癌临床分期呈负相关。生存分析结果显示，肿瘤中KCTD9低表达的患者预后更差。功能实验揭示，在体外与体内模型中，过表达KCTD9可抑制结直肠癌细胞的增殖与转移，而敲低KCTD9则会促进结直肠癌细胞的增殖与转移。通过在结直肠癌细胞中过表达或敲低KCTD9，研究发现KCTD9的表达可正向调控β-连环蛋白（β-catenin）的降解，进而抑制Wnt信号通路，并降低Wnt通路靶基因的表达。机制层面上，本研究发现KCTD9与β-连环蛋白介导的基因转录共激活因子ZNT9（锌转运蛋白9, Zinc Transporter 9）存在相互作用。在结直肠癌细胞中过表达KCTD9或敲低ZNT9，可增强β-连环蛋白的多泛素化修饰与蛋白酶体降解。反之，KCTD9与ZNT9的相互作用会破坏β-连环蛋白与ZNT9的结合，从而降低β-连环蛋白靶基因的表达并抑制Wnt信号通路。综上，本研究结果表明KCTD9作为一种肿瘤抑制因子，通过失活Wnt/β-连环蛋白通路抑制结直肠癌细胞的增殖与转移。此外，KCTD9在结直肠癌中频繁下调，提示其可作为结直肠癌潜在的预后与治疗靶点。针对邻近正常结肠黏膜、原发性结肠腺癌及远处肝转移灶的RNA测序（RNA-seq）数据开展的比较基因表达谱分析