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Methylated RNA Immunoprecipitation Assay to Study m5C Modification in Arabidopsis

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

Wen Wang¹, Hai-Tong Wang¹, Yang Guo¹

¹School of Pharmacy, XinXiang Medical University, Xinxiang 453003, Henan, China; International Joint Laboratory of Recombinant Drug Protein Expression System, Xinxiang 453003, Henan, China; Henan Engineering Research Center for Biopharmaceutical Innovation, Xinxiang Medical University, Xinxiang 453003, Henan, China.

International Journal of Biological Macromolecules

|April 27, 2025

Summary

N6-methyladenosine (m6A) RNA modification regulates therapeutic protein production in Chinese hamster ovary (CHO) cells. Inhibiting specific genes enhances protein yield, offering strategies for cell line optimization.

Keywords:

Chinese hamster ovary (CHO) cells MeRIP-seq RNA-seq Recombinant protein expression m6A modification

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A Method for Measuring RNA N6-methyladenosine Modifications in Cells and Tissues

A Method for Measuring RNA N6-methyladenosine Modifications in Cells and Tissues

Published on: December 5, 2016

Area of Science:

Biotechnology
Molecular Biology
Cell Engineering

Background:

Chinese hamster ovary (CHO) cells are crucial for recombinant therapeutic protein production.
Enhancing transgene expression and production stability in CHO cells is a persistent challenge.
The role of N6-methyladenosine (m6A) RNA modification in recombinant protein expression is largely unexplored.

Purpose of the Study:

To investigate the role of m6A RNA modification in recombinant protein production in CHO cells.
To identify m6A-related regulatory mechanisms impacting protein expression levels.
To explore strategies for optimizing CHO cell lines for enhanced protein yield.

Main Methods:

m6A-specific methylated RNA immunoprecipitation sequencing (MeRIP-seq) was performed on high- and low-producing recombinant adalimumab (ADM) CHO cell lines.
Differential methylation peaks were identified and correlated with heavy chain (HC)/light chain (LC) expression.
Gene expression analysis and pharmacological inhibition of key factors were conducted.

Main Results:

668 differentially methylated peaks were identified between high- and low-producing cell lines.
m6A methylation patterns positively correlated with HC/LC expression levels.
Inhibition of Gli2 or Met enhanced ADM production and suppressed target gene expression, implicating PI3K-Akt and Hippo signaling pathways.

Conclusions:

m6A RNA modification plays a functional role in regulating recombinant protein production in CHO cells.
m6A-mediated regulation involves factors like Igf2bp2, Gli2, and Met, potentially through PI3K-Akt and Hippo signaling.
Pharmacological targeting of Gli2 or Met offers a viable strategy for CHO cell line optimization and enhanced protein yield.