环状RNA circ_0001591通过靶向miR-20a-3p和miR-34a-5p并借助AXL和FRA1蛋白促进黑色素瘤细胞迁移。

Circular RNA circ_0001591 Contributes to Melanoma Cell Migration Through AXL and FRA1 Proteins by Targeting miR-20a-3p and miR-34a-5p.

作者信息

Orlandi Elisa, De Tomi Elisa, Belpinati Francesca, Menegazzi Marta, Gomez-Lira Macarena, Romanelli Maria Grazia, Trabetti Elisabetta

机构信息

Section of Biology and Genetics, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Strada Le Grazie, 8, 37134 Verona, Italy.

Section of Biological Chemistry, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Strada Le Grazie, 8, 37134 Verona, Italy.

出版信息

Genes (Basel). 2025 Jul 30;16(8):921. doi: 10.3390/genes16080921.

Abstract

Different risk factors are involved in the initiation and progression of melanoma. In particular, genetic and epigenetic pathways are involved in all stages of melanoma and are exploited in therapeutic approaches. This study investigated the role of circular RNA circ_0001591 in melanoma cell migration. Three different melanoma cell lines were transfected with siRNA targeting circ_0001591 and with mimic or inhibitor molecules for miR-20a-3p and miR-34a-5p. Gene and protein expression levels were analyzed by RT-qPCR and Western blot, respectively. Dual luciferase reporter assays were performed to confirm the direct interaction of miR-20a-3p and miR-34a-5p with circ_0001591, as well as with the 3'UTRs of AXL (for both miRNAs) and (miR-34a-5p only). Wound healing assays were conducted to assess cell migration velocity. The silencing of circ_0001591 significantly reduces the migration ability of melanoma cell lines. This downregulation was associated with an increased expression of miR-20a-3p and miR-34a-5p. Dual luciferase reporter assays confirmed the direct binding of both miRNAs to circ_0001591, supporting its role as a molecular sponge. The same assays also verified that miR-20a-3p directly targets the 3'UTR of , while miR-34a-5p binds the 3'UTRs of both and . Western blot analysis showed that the modulation of this axis affects the expression levels of the AXL and FRA1 oncoproteins. Our findings demonstrate that circ_0001591 promotes melanoma migration by sponging miR-20a-3p and miR-34a-5p, thereby indirectly modulating the expression of AXL and FRA1 oncoprotein. Further investigations of this new regulatory network are needed to better understand its role in melanoma progression and to support the development of targeted therapies.

摘要

不同的风险因素参与黑色素瘤的起始和进展。特别是,基因和表观遗传途径参与黑色素瘤的各个阶段,并在治疗方法中得到应用。本研究调查了环状RNA circ_0001591在黑色素瘤细胞迁移中的作用。用靶向circ_0001591的小干扰RNA(siRNA)以及miR-20a-3p和miR-34a-5p的模拟物或抑制剂分子转染三种不同的黑色素瘤细胞系。分别通过逆转录定量聚合酶链反应(RT-qPCR)和蛋白质免疫印迹法分析基因和蛋白质表达水平。进行双荧光素酶报告基因测定以确认miR-20a-3p和miR-34a-5p与circ_0001591以及AXL的3'非翻译区(3'UTR)(两种miRNA均涉及)和FRA1(仅miR-34a-5p涉及)的直接相互作用。进行伤口愈合试验以评估细胞迁移速度。circ_0001591的沉默显著降低黑色素瘤细胞系的迁移能力。这种下调与miR-20a-3p和miR-34a-5p表达增加有关。双荧光素酶报告基因测定证实两种miRNA均与circ_0001591直接结合,支持其作为分子海绵的作用。相同的测定还验证了miR-20a-3p直接靶向FRA1的3'UTR,而miR-34a-5p与AXL和FRA1的3'UTR均结合。蛋白质免疫印迹分析表明该轴的调节影响AXL和FRA1癌蛋白的表达水平。我们的研究结果表明,circ_0001591通过吸附miR-20a-3p和miR-34a-5p促进黑色素瘤迁移,从而间接调节AXL和FRA1癌蛋白的表达。需要对这个新的调控网络进行进一步研究,以更好地了解其在黑色素瘤进展中的作用,并支持靶向治疗的开发。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a040/12385530/3dc31cecdf6a/genes-16-00921-g001.jpg

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