MITF (rs7623610)和CREB1 (rs10932201)基因的内含子变异可能提高人类黑色素瘤细胞系的剪接效率

IF 1.5 4区医学 Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis Pub Date : 2021-07-01 DOI:10.1016/j.mrfmmm.2021.111763

Juliana Carron , Caroline Torricelli , Janet Keller Silva , Lilian de Oliveira Coser , Carmen Silvia Passos Lima , Gustavo Jacob Lourenço

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引用次数: 0

摘要

我们之前报道了MITF (c.938−325G>A, rs7623610)和CREB1 (c.303+373G>A, rs10932201)基因的内含子单核苷酸变异(snv)与皮肤黑色素瘤(CM)的风险、侵袭性和预后相关。在本研究中，我们研究了上述snv对剪接模式和效率的影响。我们利用MITF和CREB1的野生型和变异等位基因构建了迷你基因，以评估snv对剪接的影响。将这些基因转染到人黑色素瘤细胞系SK-MEL-28中。RT-PCR和DNA测序研究了这些构建体的剪接模式。采用qPCR检测Minigenes构建体的剪接效率及HNRNPA1和SF1剪接基因的表达。我们发现MITF和CREB1 snv不改变剪接模式，但影响剪接效率。MITF-A (p= 0.03)和CREB1-A (p= 0.005)变异基因产生的mRNA增加。此外，HNRNPA1和SF1的mRNA水平在变异基因MITF-A (p= 0.04)和CREB1-A (p= 0.005)中也较低。我们首次描述了MITF rs7623610和CREB1 rs10932201 snv在剪接效率中的潜在重要性及其与CM的关系。

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Intronic variants of MITF (rs7623610) and CREB1 (rs10932201) genes may enhance splicing efficiency in human melanoma cell line

We previously reported that intronic single nucleotide variations (SNVs) in MITF (c.938−325G>A, rs7623610) and CREB1 (c.303+373G>A, rs10932201) genes were associated with risk, aggressiveness, and prognosis of cutaneous melanoma (CM). In this study, we investigated the influence of the above SNVs in splicing patterns and efficiency. We constructed minigenes with wild type and variant alleles from MITF and CREB1 to assess the effect of the SNVs on splicing. The minigenes were transfected in the human melanoma cell line (SK-MEL-28). RT-PCR and DNA sequencing investigated the constructs’ splicing patterns. Minigenes constructs’ splicing efficiency and HNRNPA1 and SF1 splicing genes’ expression were investigated by qPCR. We found that MITF and CREB1 SNVs did not alter the splicing pattern, but they influenced the splicing efficiency. MITF-A (p= 0.03) and CREB1-A (p= 0.005) variant minigenes yielded an increase of mRNA generated from the constructions. Additionally, lower mRNA levels of HNRNPA1 and SF1 were seen in the variant minigenes MITF-A (p= 0.04) and CREB1-A (p= 0.005). We described for the first time the potential importance of MITF rs7623610 and CREB1 rs10932201 SNVs in splicing efficiency and its relationship with CM.

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来源期刊

Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis 生物-毒理学

CiteScore

4.90

自引率

0.00%

发文量

审稿时长

51 days

期刊介绍： Mutation Research (MR) provides a platform for publishing all aspects of DNA mutations and epimutations, from basic evolutionary aspects to translational applications in genetic and epigenetic diagnostics and therapy. Mutations are defined as all possible alterations in DNA sequence and sequence organization, from point mutations to genome structural variation, chromosomal aberrations and aneuploidy. Epimutations are defined as alterations in the epigenome, i.e., changes in DNA methylation, histone modification and small regulatory RNAs. MR publishes articles in the following areas: Of special interest are basic mechanisms through which DNA damage and mutations impact development and differentiation, stem cell biology and cell fate in general, including various forms of cell death and cellular senescence. The study of genome instability in human molecular epidemiology and in relation to complex phenotypes, such as human disease, is considered a growing area of importance. Mechanisms of (epi)mutation induction, for example, during DNA repair, replication or recombination; novel methods of (epi)mutation detection, with a focus on ultra-high-throughput sequencing. Landscape of somatic mutations and epimutations in cancer and aging. Role of de novo mutations in human disease and aging; mutations in population genomics. Interactions between mutations and epimutations. The role of epimutations in chromatin structure and function. Mitochondrial DNA mutations and their consequences in terms of human disease and aging. Novel ways to generate mutations and epimutations in cell lines and animal models.