a Department of Neurosurgery , The Second Affiliated Hospital, Fujian Medical University , Quanzhou , PR China.
b Department of Neurosurgery , Affiliated Hospital of North Sichuan Medical College , Nanchong , PR China.
Artif Cells Nanomed Biotechnol. 2018;46(sup1):608-615. doi: 10.1080/21691401.2018.1431654. Epub 2018 Jan 29.
This study aims to via unveiling the novel mechanisms of KLF16 in regulating expression of genes involved in glioma.
KLF16 or KLF16-siRNA was transfected to U87MG cells by lentivirus. Colony formation assay was applied for detecting cell proliferation. MTT assay was adopted to assess cell viability. TUNEL assay was selected to evaluate cell apoptosis. Flow cytometry was used to determine cell cycle. Real-time PCR was performed to test mRNA expression. Western blot was used to detect protein level. Luciferase assay was applied to confirm the regulatory relationship between KLF16 and Mitochondrial transcription factor A (TFAM). Chromatin immunoprecipitation was adopted to test the protein binding site. The nude mouse transplantation tumour experiment was selected to test cancer cell proliferation in vivo.
KLF16 was decreased in glioma cells and tissues. KLF16 obviously restrained U87MG cell proliferation both in vivo and in vitro. KLF16 transfection reduced mRNA and protein levels related to cell proliferation. KLF16 targeted a putative binding site near the transcription start sites (TSSs) of TFAM gene, thus suppressing glioma cell proliferation. KLF16-siRNA exhibited the opposite impact. KLF16 presented significant negative correlation with TFAM level in glioma patients.
KLF16 is a key regulator of glioma cell proliferation by directly targeting TFAM.
本研究旨在揭示 KLF16 调控胶质瘤相关基因表达的新机制。
通过慢病毒转染将 KLF16 或 KLF16-siRNA 转染至 U87MG 细胞。集落形成实验用于检测细胞增殖。MTT 法评估细胞活力。TUNEL 法检测细胞凋亡。流式细胞术检测细胞周期。实时 PCR 检测 mRNA 表达。Western blot 检测蛋白水平。荧光素酶报告基因检测 KLF16 与线粒体转录因子 A（TFAM）的调控关系。染色质免疫沉淀法检测蛋白结合位点。裸鼠移植瘤实验检测体内肿瘤细胞增殖。
KLF16 在胶质瘤细胞和组织中表达降低。KLF16 明显抑制 U87MG 细胞在体内和体外的增殖。KLF16 转染降低与细胞增殖相关的 mRNA 和蛋白水平。KLF16 靶向 TFAM 基因转录起始位点（TSS）附近的一个假定结合位点，从而抑制胶质瘤细胞增殖。KLF16-siRNA 则表现出相反的影响。KLF16 在胶质瘤患者中的表达与 TFAM 水平呈显著负相关。
KLF16 通过直接靶向 TFAM 成为胶质瘤细胞增殖的关键调节因子。
