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Abstract：
BACKGROUND: Circular RNAs (circRNAs) have been shown to be involved in tumorigenesis and progression. However, the role of circGLIS3 (hsa_circ_0002874) in prostate cancer (PCa) has yet not been reported.
METHODS: Candidate circRNA were determined through comprehensive analysis of public datasets, PCa cell lines, and tissues data. A series of cellular functional assays, including CCK-8, colony formation, wound healing, and transwell assays were performed. Subsequently, RNA sequencing, RNA immunoprecipitation, methylated RNA immunoprecipitation, microRNA pulldown, luciferase reporter assay, and western blot were used to explore the underlying molecular mechanisms. Moreover, the xenograft tumor mouse model was established to elucidate the function of circGLIS3.
RESULTS: CircGLIS3, derived from exon 2 of the parental GLIS3 gene, was identified as a novel oncogenic circRNA in PCa that was closely associated with the biochemical recurrence. Its expression levels were upregulated in PCa tissues and cell lines as well as enzalutamide high-resistant cells. The cellular functional assays revealed that circGLIS3 promoted PCa cell proliferation, migration, and invasion. METTL3-mediated N6-methyladenosine (m6A) modification maintained its upregulation by enhancing its stability. Mechanically, CircGLIS3 sponged miR-661 to upregulate MDM2, thus regulating the p53 signaling pathway to promote cell proliferation, migration, and invasion. Furthermore, in vitro and in vivo experiments, the knockdown of circGLIS3 improved the response of PCa cells to ARSI therapies such as enzalutamide.
CONCLUSIONS: METTL3-mediated m6A modification of circGLIS3 regulates the p53 signaling pathway via the miR-661/MDM2 axis, thereby facilitating PCa progression. Meanwhile, this study unveils a promising potential target for ARSI therapy for PCa.
METHODS: Candidate circRNA were determined through comprehensive analysis of public datasets, PCa cell lines, and tissues data. A series of cellular functional assays, including CCK-8, colony formation, wound healing, and transwell assays were performed. Subsequently, RNA sequencing, RNA immunoprecipitation, methylated RNA immunoprecipitation, microRNA pulldown, luciferase reporter assay, and western blot were used to explore the underlying molecular mechanisms. Moreover, the xenograft tumor mouse model was established to elucidate the function of circGLIS3.
RESULTS: CircGLIS3, derived from exon 2 of the parental GLIS3 gene, was identified as a novel oncogenic circRNA in PCa that was closely associated with the biochemical recurrence. Its expression levels were upregulated in PCa tissues and cell lines as well as enzalutamide high-resistant cells. The cellular functional assays revealed that circGLIS3 promoted PCa cell proliferation, migration, and invasion. METTL3-mediated N6-methyladenosine (m6A) modification maintained its upregulation by enhancing its stability. Mechanically, CircGLIS3 sponged miR-661 to upregulate MDM2, thus regulating the p53 signaling pathway to promote cell proliferation, migration, and invasion. Furthermore, in vitro and in vivo experiments, the knockdown of circGLIS3 improved the response of PCa cells to ARSI therapies such as enzalutamide.
CONCLUSIONS: METTL3-mediated m6A modification of circGLIS3 regulates the p53 signaling pathway via the miR-661/MDM2 axis, thereby facilitating PCa progression. Meanwhile, this study unveils a promising potential target for ARSI therapy for PCa.
摘要：
背景：环状RNA（circularRNAs，circRNAs）已被证明与肿瘤发生和发展有关。然而,circGLIS3(hsa_circ_0002874)在前列腺癌(PCa)中的作用尚未见报道。
方法：通过对公共数据集的综合分析确定候选circRNA，PCa细胞系,和组织数据。一系列的细胞功能检测,包括CCK-8，集落形成，伤口愈合，和transwell测定进行。随后，RNA测序，RNA免疫沉淀，甲基化RNA免疫沉淀，microRNA下拉，荧光素酶报告分析，和蛋白质印迹用于探索潜在的分子机制。此外，建立异种移植瘤小鼠模型以阐明circGLIS3的功能。
结果：CircGLIS3，来自亲本GLIS3基因的外显子2，在PCa中被鉴定为与生化复发密切相关的新型致癌circRNA。其表达水平在PCa组织和细胞系以及恩杂鲁胺高抗性细胞中上调。细胞功能测定显示，circGLIS3促进PCa细胞增殖，迁移,和入侵。METTL3介导的N6-甲基腺苷（m6A）修饰通过增强其稳定性来维持其上调。机械上,CircGLIS3激活miR-661上调MDM2,从而调节p53信号通路促进细胞增殖,迁移,和入侵。此外，体外和体内实验，circGLIS3的敲减改善了PCa细胞对ARSI治疗如恩杂鲁胺的反应.
结论：METTL3介导的m6A修饰circGLIS3通过miR-661/MDM2轴调节p53信号通路，从而促进PCa进展。同时,这项研究揭示了ARSI治疗PCa的一个有希望的潜在靶点.
方法：通过对公共数据集的综合分析确定候选circRNA，PCa细胞系,和组织数据。一系列的细胞功能检测,包括CCK-8，集落形成，伤口愈合，和transwell测定进行。随后，RNA测序，RNA免疫沉淀，甲基化RNA免疫沉淀，microRNA下拉，荧光素酶报告分析，和蛋白质印迹用于探索潜在的分子机制。此外，建立异种移植瘤小鼠模型以阐明circGLIS3的功能。
结果：CircGLIS3，来自亲本GLIS3基因的外显子2，在PCa中被鉴定为与生化复发密切相关的新型致癌circRNA。其表达水平在PCa组织和细胞系以及恩杂鲁胺高抗性细胞中上调。细胞功能测定显示，circGLIS3促进PCa细胞增殖，迁移,和入侵。METTL3介导的N6-甲基腺苷（m6A）修饰通过增强其稳定性来维持其上调。机械上,CircGLIS3激活miR-661上调MDM2,从而调节p53信号通路促进细胞增殖,迁移,和入侵。此外，体外和体内实验，circGLIS3的敲减改善了PCa细胞对ARSI治疗如恩杂鲁胺的反应.
结论：METTL3介导的m6A修饰circGLIS3通过miR-661/MDM2轴调节p53信号通路，从而促进PCa进展。同时,这项研究揭示了ARSI治疗PCa的一个有希望的潜在靶点.
