College of Pharmacy and Medical Research Center, Chungbuk National University, Osongsaengmyeong 1-ro 194-21, Osong-eup, Heungduk-gu, Cheongju, Chungbuk, 28160, Republic of Korea.
Department of Pharmacy, Wonkwang University, 460 Iksandae-ro, Iksan, Jeonbuk, 54538, Republic of Korea.
Theranostics. 2018 Aug 7;8(16):4409-4428. doi: 10.7150/thno.26467. eCollection 2018.
Chitinase 3-like 1 (Chi3L1) protein is up-regulated in various diseases including solid cancers. According to Genome-Wide Association Study (GWAS)/Online Mendelian Inheritance in Man (OMIM)/Differentially Expressed Gene (DEG) analyses, Chi3L1 is associated with 38 cancers, and more highly associated with cancer compared to other oncogenes such as EGFR, TNFα, etc. However, the mechanisms and pathways by which Chi3L1 is associated with cancer are not clear. In current study, we investigated the role of Chi3L1 in lung metastasis. We performed the differentially expressed gene analysis to explore the genes which are associated with Chi3L1 using the web-based platform from Biomart. We investigated the metastases in lung tissues of C57BL/6 mice injected with B16F10 melanoma following treatment with Ad-shChi3L1. We also investigated the expression of USF1 and Chi3L1 in Chi3L1 KD mice lung tissues by Western blotting and IHC. We also analyzed lung cancer cells metastases induced by Chi3L1 using migration and cell proliferation assay in human lung cancer cell lines. The involvement of miR-125a-3p in Chi3L1 regulation was determined by miRNA qPCR and luciferase reporter assay. We showed that melanoma metastasis in lung tissues was significantly reduced in Chi3L1 knock-down mice, accompanied by down-regulation of MMP-9, MMP-13, VEGF, and PCNA in Chi3L1 knock-down mice lung tissue, as well as in human lung cancer cell lines. We also found that USF1 was conversely expressed against Chi3L1. USF1 was increased by knock-down of Chi3L1 in mice lung tissues, as well as in human lung cancer cell lines. In addition, knock-down of USF1 increased Chi3L1 levels in addition to augmenting metastasis cell migration and proliferation in mice model, as well as in human cancer cell lines. Moreover, in human lung tumor tissues, the expression of Chi3L1 was increased but USF1 was decreased in a stage-dependent manner. Finally, Chi3L1 expression was strongly regulated by the indirect translational suppressing activity of USF1 through induction of miR-125a-3p, a target of Chi3L1. Metastases in mice lung tissues and human lung cancer cell lines were decreased by KD of Chi3L1. USF1 bound to the Chi3L1 promoter, however, Chi3L1 expression was decreased by USF1, despite USF1 enhancing the transcriptional activity of Chi3L1. We found that USF1 induced miR-125a-3p levels which suppressed Chi3L1 expression. Ultimately, our results suggest that lung metastasis is suppressed by knock-down of Chi3L1 through miR-125a-3p-mediated up-regulation of USF1.
几丁质酶 3 样蛋白 1（Chi3L1）在包括实体瘤在内的各种疾病中呈上调表达。根据全基因组关联研究（GWAS）/在线孟德尔遗传数据库（OMIM）/差异表达基因（DEG）分析，Chi3L1 与 38 种癌症相关，与 EGFR、TNFα 等其他癌基因相比，与癌症的相关性更高。然而，Chi3L1 与癌症相关的机制和途径尚不清楚。在本研究中，我们研究了 Chi3L1 在肺转移中的作用。我们使用基于网络的 Biomart 平台进行差异表达基因分析，以探讨与 Chi3L1 相关的基因。我们研究了用 Ad-shChi3L1 处理后 B16F10 黑色素瘤注射的 C57BL/6 小鼠肺组织中的转移。我们还通过 Western blot 和 IHC 研究了 Chi3L1 KD 小鼠肺组织中 USF1 和 Chi3L1 的表达。我们还使用人肺癌细胞系中的迁移和细胞增殖测定分析了 Chi3L1 诱导的肺癌细胞转移。通过 miRNA qPCR 和荧光素酶报告基因测定确定了 miR-125a-3p 在 Chi3L1 调节中的作用。我们表明，Chi3L1 敲低小鼠的黑色素瘤肺转移明显减少，伴随 Chi3L1 敲低小鼠肺组织中 MMP-9、MMP-13、VEGF 和 PCNA 的下调，以及人肺癌细胞系中的下调。我们还发现 USF1 与 Chi3L1 呈反式表达。在小鼠肺组织和人肺癌细胞系中，Chi3L1 的敲低增加了 USF1 的表达。此外，USF1 的敲低除了增强小鼠模型和人癌细胞系中的转移细胞迁移和增殖外，还增加了 Chi3L1 水平。此外，在人肺肿瘤组织中，Chi3L1 的表达呈阶段依赖性增加，而 USF1 减少。最后，Chi3L1 的表达受 USF1 通过诱导 Chi3L1 的靶标 miR-125a-3p 的间接翻译抑制活性的强烈调节。Chi3L1 的 KD 降低了小鼠肺组织和人肺癌细胞系中的转移。USF1 与 Chi3L1 启动子结合，但 Chi3L1 的表达被 USF1 降低，尽管 USF1 增强了 Chi3L1 的转录活性。我们发现 USF1 诱导了 miR-125a-3p 水平，从而抑制了 Chi3L1 的表达。最终，我们的结果表明，通过 miR-125a-3p 介导的 USF1 上调，Chi3L1 的敲低抑制了肺转移。
