RNASeq highlights ATF6 pathway regulators for CHO cell engineering with different impacts of ATF6β and WFS1 knockdown on fed-batch production of IgG₁

Dyllan Rives ¹, Caroline Peak ¹, Mark A Blenner ^2,3

⁰Department of Chemical & Biomolecular Engineering, Clemson University, 206 S. Palmetto Blvd., Clemson, SC, 29634-0909, USA.

Scientific Reports +

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June 19, 2024

View abstract on PubMed

Summary

Optimizing the unfolded protein response (UPR) in Chinese hamster ovary (CHO) cells enhances therapeutic protein production. Knocking down ATF6β improved UPR and productivity, while WFS1 knockdown decreased protein titer.

Area Of Science

Biotechnology
Cell Biology
Molecular Biology

Background

Endoplasmic reticulum (ER) stress from high protein secretion challenges Chinese hamster ovary (CHO) cell biomanufacturing.
The unfolded protein response (UPR) is crucial for maintaining ER homeostasis and optimizing therapeutic protein production.

Purpose Of The Study

To investigate the ER stress response and UPR activation in an immunoglobulin G1 (IgG1) producing CHO cell line compared to its parental host cell line.
To identify potential engineering targets within the UPR pathway to enhance biomanufacturing productivity.

Main Methods

Fed-batch cell culture of IgG1-producing and host CHO cell lines.
High-throughput RNA sequencing (RNASeq) and quantitative polymerase chain reaction (qPCR) for differential gene expression analysis.
Gene knockdown experiments targeting ATF6β and WFS1.

Main Results

The IgG1 producer exhibited UPR activation compared to the host cell line, with transient upregulation of ATF6α target mRNAs.
Knockdown of ATF6β decreased cell growth but maintained titer, while WFS1 knockdown decreased titer without affecting growth.
ATF6β knockdown improved later-stage UPR activation and overall productivity, whereas WFS1 knockdown negatively impacted UPR and product mRNA expression.

Conclusions

ATF6β and WFS1 are identified as key regulators of the ATF6 arm of the UPR in CHO cells.
Targeting ATF6β presents a viable strategy for enhancing UPR and improving therapeutic protein productivity in CHO cell biomanufacturing.
WFS1 appears to play a distinct role, potentially limiting productivity under certain conditions.