Department of Anatomical and Cellular Pathology, State Key Laboratory in Oncology in South China, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong SAR, China.
Institute of Digestive Disease, Partner State Key Laboratory of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, China.
Oncogene. 2018 Mar;37(9):1159-1174. doi: 10.1038/s41388-017-0029-7. Epub 2017 Dec 13.
Slit-Robo GTPase-activating protein 1 (SRGAP1) functions as a GAP for Rho-family GTPases and downstream of Slit-Robo signaling. We aim to investigate the biological function of SRGAP1 and reveal its regulation by deregulated microRNAs (miRNAs) in gastric cancer (GC). mRNA and protein expression of SRGAP1 were examined by quantitative reverse transcription PCR (qRT-PCR) and western blot. The biological role of SRGAP1 was demonstrated through siRNA-mediated knockdown experiments. The regulation of SRGAP1 by miR-340 and miR-124 was confirmed by western blot, dual luciferase activity assays and rescue experiments. SRGAP1 is overexpressed in 9 out of 12 (75.0%) GC cell lines. In primary GC samples from TCGA cohort, SRGAP1 shows gene amplification in 5/258 (1.9%) of cases and its mRNA expression demonstrates a positive correlation with copy number gain. Knockdown of SRGAP1 in GC cells suppressed cell proliferation, reduced colony formation, and significantly inhibited cell invasion and migration. Luciferase reporter assays revealed that SRGAP1 knockdown significantly inhibited Wnt/β-catenin pathway. In addition, SRGAP1 was found to be a direct target of two tumor-suppressive miRNAs, miR-340 and miR-124. Concordantly, these two miRNAs were downregulated in primary gastric tumors and these decreasing levels w5ere associated with poor outcomes. Expression of miR-340 and SRGAP1 displayed a reverse relationship in primary samples and re-expressed SRGAP1, rescued the anti-cancer effects of miR-340. Taken together, these data strongly suggest that, apart from gene amplification and mutation, the activation of SRGAP1 in GC is partly due to the downregulation of tumor-suppressive miRNAs, miR-340 and miR-124. Thus SRGAP1 is overexpressed in gastric carcinogenesis and plays an oncogenic role through activating Wnt/β-catenin pathway.
Slit-Robo GTPase-activating protein 1（SRGAP1）作为 Rho 家族 GTPases 的 GAP 并在 Slit-Robo 信号下游发挥作用。我们旨在研究 SRGAP1 的生物学功能，并揭示其在胃癌（GC）中受失调的 microRNAs（miRNAs）的调控。通过定量逆转录 PCR（qRT-PCR）和 Western blot 检测 SRGAP1 的 mRNA 和蛋白表达。通过 siRNA 介导的敲低实验证明了 SRGAP1 的生物学作用。通过 Western blot、双荧光素酶活性测定和挽救实验证实了 SRGAP1 受 miR-340 和 miR-124 的调控。SRGAP1 在 12 个 GC 细胞系中的 9 个（75.0%）中过表达。在 TCGA 队列的原发性 GC 样本中，SRGAP1 在 5/258（1.9%）病例中显示基因扩增，其 mRNA 表达与拷贝数增益呈正相关。GC 细胞中 SRGAP1 的敲低抑制了细胞增殖，减少了集落形成，显著抑制了细胞侵袭和迁移。荧光素酶报告实验表明，SRGAP1 的敲低显著抑制了 Wnt/β-catenin 通路。此外，发现 SRGAP1 是两种肿瘤抑制性 miRNAs，miR-340 和 miR-124 的直接靶标。一致地，这两种 miRNAs 在原发性胃肿瘤中下调，并且这些降低的水平与不良预后相关。在原发性样本中，miR-340 和 SRGAP1 的表达呈相反关系，重新表达 SRGAP1 挽救了 miR-340 的抗癌作用。总之，这些数据强烈表明，除了基因扩增和突变之外，GC 中 SRGAP1 的激活部分归因于肿瘤抑制性 miRNAs，miR-340 和 miR-124 的下调。因此，SRGAP1 在胃发生癌中过度表达，并通过激活 Wnt/β-catenin 通路发挥致癌作用。
