Investigating the interaction of hsa-miR-193b-5p and hsa-circ-cnot11-0001 in multiple sclerosis disease through dual luciferase assay

IF 0.9 Q4 GENETICS & HEREDITY

Gene Reports Pub Date : 2025-07-08 DOI:10.1016/j.genrep.2025.102291

Amirreza Pashapouryeganeh , Elham Mohammed Khatrawi , Zohreh Pajohesh , Rosa Hosseinzadegan , Khaterehsadat Monirvaghefi , Seyed Abbas Pakmehr , Narges Pourdeilami , Hossein Gharedaghi , Sayedeh Fatemeh Sadat Madani , Ali Azarpey , Mona Torkaman Cheh , Asal Mir , Elham Ramezannezhad , Saminnaz Kazemi , Alireza Azani , Negin Saffarzadeh , Haniyeh Ghasrsaz , Asra Idani , Nour Mohammad Panahi , Nasibeh Sargazi Moghaddam , Moein Ghasemi

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引用次数: 0

Abstract

Background

Multiple sclerosis (MS) is a chronic autoimmune disorder characterized by dynamic neurodegeneration, basically affecting people between the ages of 20 and 40. Despite significant progress in MS research, little is understood about the underlying molecular mechanisms, particularly those related to epigenetic regulation. According to recent studies, non-coding RNAs (ncRNAs) are crucial epigenetic regulators that have a big impact on how multiple sclerosis develops and is controlled. Given the increasing interest in ncRNAs as biomarkers and therapeutic targets, it is more crucial than ever to look into their role in MS. Recent transcriptomic research showing that circRNAs and miRNAs express differently in neurological disorders makes this especially pertinent. In order to shed light on their possible regulatory role in MS, this study examines the expression levels of circ-cnot11-0001 and hsa-miR-193b-5p in patients and uses a dual luciferase assay to analyze their direct interaction.

Methods

The gene expression profile GSE61741 was used to select key miRNAs and circRNAs in MS. Expressions of hsa-miR-193b-5p and circ-cnot11-0001 were verified by RT-qPCR from blood samples of 30 MS patients and 30 controls. After that, we constructed pBud-EGFP plasmid with hsa-miR-193b-5p and psiCHECK-2 plasmid with circ-cnot11-0001. Their direct interaction was determined by measuring luciferase activity.

Result

Expression analysis revealed significant downregulation of hsa-miR-193b-5p and upregulation of hsa-circ-cnot11-0001 in MS patients compared to controls. Also, the results of the luciferase activity determined that the luciferase activity of the hsa-miR-193b-5p-pBud + cnot11-0001-WT group was significantly lower than that of the hsa-miR-193b-5p-pBud + cnot11-0001-MT group. So, circ-cnot11-0001 can direct interaction with hsa-miR-193b-5p.

Conclusion

The findings indicate a direct interaction between hsa-miR-193b-5p and circ-cnot11-0001 in MS. Therefore, circ-cnot11-0001 may sponge hsa-miR-193b-5p and dysregulate the expression of genes related to MS. In conclusion, the CNOT11-0001-hsa-miR-193b-5p network may serve as a combined biomarker for further investigation into MS.

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通过双荧光素酶测定研究hsa-miR-193b-5p和hsa-circ- not11-0001在多发性硬化症中的相互作用

多发性硬化症（MS）是一种以动态神经变性为特征的慢性自身免疫性疾病，主要影响20至40岁的人群。尽管MS研究取得了重大进展，但人们对其潜在的分子机制，特别是与表观遗传调控有关的机制知之甚少。根据最近的研究，非编码rna （ncRNAs）是重要的表观遗传调控因子，对多发性硬化症的发展和控制有重要影响。鉴于人们对ncRNAs作为生物标志物和治疗靶点的兴趣日益浓厚，研究它们在ms中的作用比以往任何时候都更加重要。最近的转录组学研究表明，circRNAs和miRNAs在神经系统疾病中的表达不同，这一点尤为重要。为了阐明它们在MS中可能的调节作用，本研究检测了circ- conn11 -0001和hsa-miR-193b-5p在患者中的表达水平，并使用双荧光素酶测定来分析它们的直接相互作用。方法采用基因表达谱GSE61741筛选MS中的关键mirna和circrna，采用RT-qPCR方法对30例MS患者和30例对照组血液样本中hsa-miR-193b-5p和circ- connot11 -0001的表达进行验证。之后，我们用hsa-miR-193b-5p构建了pbu - egfp质粒，用circ- connot11 -0001构建了psiCHECK-2质粒。它们的直接相互作用是通过测量荧光素酶活性来确定的。结果表达分析显示，与对照组相比，MS患者中hsa-miR-193b-5p显著下调，hsa-circ- connot11 -0001上调。此外，荧光素酶活性测定结果表明，hsa-miR-193b-5p-pBud + cnot11-0001-WT组的荧光素酶活性显著低于hsa-miR-193b-5p-pBud + cnot11-0001-MT组。因此circ- connot11 -0001可以直接与hsa-miR-193b-5p相互作用。结论研究结果表明，MS中hsa-miR-193b-5p与circ- cnnot11 -0001之间存在直接相互作用，因此circ- cnnot11 -0001可能会吞噬hsa-miR-193b-5p，并使MS相关基因的表达失调。综上所述，cnnot11 -0001-hsa-miR-193b-5p网络可能作为MS进一步研究的联合生物标志物。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Gene Reports Biochemistry, Genetics and Molecular Biology-Genetics

CiteScore

3.30

自引率

7.70%

发文量

246

审稿时长

49 days

期刊介绍： Gene Reports publishes papers that focus on the regulation, expression, function and evolution of genes in all biological contexts, including all prokaryotic and eukaryotic organisms, as well as viruses. Gene Reports strives to be a very diverse journal and topics in all fields will be considered for publication. Although not limited to the following, some general topics include: DNA Organization, Replication & Evolution -Focus on genomic DNA (chromosomal organization, comparative genomics, DNA replication, DNA repair, mobile DNA, mitochondrial DNA, chloroplast DNA). Expression & Function - Focus on functional RNAs (microRNAs, tRNAs, rRNAs, mRNA splicing, alternative polyadenylation) Regulation - Focus on processes that mediate gene-read out (epigenetics, chromatin, histone code, transcription, translation, protein degradation). Cell Signaling - Focus on mechanisms that control information flow into the nucleus to control gene expression (kinase and phosphatase pathways controlled by extra-cellular ligands, Wnt, Notch, TGFbeta/BMPs, FGFs, IGFs etc.) Profiling of gene expression and genetic variation - Focus on high throughput approaches (e.g., DeepSeq, ChIP-Seq, Affymetrix microarrays, proteomics) that define gene regulatory circuitry, molecular pathways and protein/protein networks. Genetics - Focus on development in model organisms (e.g., mouse, frog, fruit fly, worm), human genetic variation, population genetics, as well as agricultural and veterinary genetics. Molecular Pathology & Regenerative Medicine - Focus on the deregulation of molecular processes in human diseases and mechanisms supporting regeneration of tissues through pluripotent or multipotent stem cells.