m6A modification profiles of the CHO cells with differential recombinant protein expression using MeRIP-seq/RNA-seq

IF 7.7 1区化学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

International Journal of Biological Macromolecules Pub Date : 2025-05-01 DOI:10.1016/j.ijbiomac.2025.143429

Wen Wang , Hai-Tong Wang , Yang Guo , Qi Zhao , Jiang-Tao Lu , Zhao-Ming Cui , Xi Zhang , Le-Le Qiu , Xiao-Yin Wang , Tian-Yun Wang , Yan-Long Jia

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引用次数: 0

Abstract

Chinese hamster ovary (CHO) cells remain the primary host system for recombinant therapeutic protein production. Enhancing transgene expression efficiency while maintaining stable production persists as a key challenge in CHO cell engineering. While N6-methyladenosine (m6A) modification – the most abundant RNA methylation – regulates RNA stability and translational efficiency, its role in modulating recombinant protein expression remains underexplored. In this study, through m6A-specific methylated RNA immunoprecipitation sequencing (MeRIP-seq) of high- (ADM-H) and low- (ADM-L) recombinant adalimumab (ADM)-producing CHO cell lines, we identified 668 differentially methylated peaks. Notably, m6A methylation patterns showed positive correlation with heavy chain (HC)/light chain (LC) expression levels between ADM-H and ADM-L cell lines. Differential expression of factors, such as Igf2bp2, Gli2, and Met correlated with PI3K-Akt and Hippo signaling pathways, suggesting m6A-mediated regulatory functions of recombinant protein expression in CHO cells. Furthermore, pharmacological inhibition of Gli2 or Met in cell culture effectively enhanced ADM production while suppressing target gene expression. These findings elucidate m6A's functional role in recombinant protein production and provide actionable strategies for CHO cell line optimization.

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利用MeRIP-seq/RNA-seq技术对具有差异重组蛋白表达的CHO细胞进行m6A修饰谱分析

中国仓鼠卵巢（CHO）细胞仍然是重组治疗性蛋白生产的主要宿主系统。提高转基因表达效率的同时保持稳定的产量一直是CHO细胞工程的关键挑战。虽然n6 -甲基腺苷（m6A）修饰-最丰富的RNA甲基化-调节RNA稳定性和翻译效率，但其在调节重组蛋白表达中的作用仍未得到充分研究。在这项研究中，通过对高（ADM- h）和低（ADM- l）重组阿达木单抗（ADM）产生的CHO细胞系进行m6a特异性甲基化RNA免疫沉淀测序（MeRIP-seq），我们鉴定了668个差异甲基化峰。值得注意的是，在ADM-H和ADM-L细胞系中，m6A甲基化模式与重链(HC)/轻链（LC）表达水平呈正相关。Igf2bp2、Gli2和Met等因子的差异表达与PI3K-Akt和Hippo信号通路相关，提示m6a介导的重组蛋白在CHO细胞中的表达调节功能。此外，在细胞培养中抑制Gli2或Met可有效提高ADM的产生，同时抑制靶基因的表达。这些发现阐明了m6A在重组蛋白生产中的功能作用，并为CHO细胞系优化提供了可行的策略。

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

International Journal of Biological Macromolecules 生物-生化与分子生物学

CiteScore

13.70

自引率

9.80%

发文量

2728

审稿时长

64 days

期刊介绍： The International Journal of Biological Macromolecules is a well-established international journal dedicated to research on the chemical and biological aspects of natural macromolecules. Focusing on proteins, macromolecular carbohydrates, glycoproteins, proteoglycans, lignins, biological poly-acids, and nucleic acids, the journal presents the latest findings in molecular structure, properties, biological activities, interactions, modifications, and functional properties. Papers must offer new and novel insights, encompassing related model systems, structural conformational studies, theoretical developments, and analytical techniques. Each paper is required to primarily focus on at least one named biological macromolecule, reflected in the title, abstract, and text.