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Dynamics of unfolded protein response in recombinant CHO cells

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Abstract

Genes in the protein secretion pathway have been targeted to increase productivity of monoclonal antibodies in Chinese hamster ovary cells. The results have been highly variable depending on the cell type and the relative amount of recombinant and target proteins. This paper presents a comprehensive study encompassing major components of the protein processing pathway in the endoplasmic reticulum (ER) to elucidate its role in recombinant cells. mRNA profiles of all major ER chaperones and unfolded protein response (UPR) pathway genes are measured at a series of time points in a high-producing cell line under the dynamic environment of a batch culture. An initial increase in IgG heavy chain mRNA levels correlates with an increase in productivity. We observe a parallel increase in the expression levels of majority of chaperones. The chaperone levels continue to increase until the end of the batch culture. In contrast, calreticulin and ERO1-l alpha, two of the lowest expressed genes exhibit transient time profiles, with peak induction on day 3. In response to increased ER stress, both the GCN2/PKR-like ER kinase and inositol-requiring enzyme-1alpha (Ire1α) signalling branch of the UPR are upregulated. Interestingly, spliced X-Box binding protein 1 (XBP1s) transcription factor from Ire1α pathway is detected from the beginning of the batch culture. Comparison with the expression levels in a low producer, show much lower induction at the end of the exponential growth phase. Thus, the unfolded protein response strongly correlates with the magnitude and timing of stress in the course of the batch culture.

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Acknowledgments

This work was partially supported by a grant from Department of Biotechnology, Government of India. We would like to thank Dr. Miranda Yap and Dr. Niki Wong, Bioprocessing Technology Institute, Singapore for providing the CHO cell lines. We would also like to thank Prof. Wei Shou Hu of University of Minnesota for useful discussions and comments on the manuscript.

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  1. Department of Chemical Engineering, Indian Institute of Technology Bombay, Powai, Mumbai, 400076, India

    Kamal Prashad & Sarika Mehra

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  1. Kamal Prashad
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Correspondence to Sarika Mehra.

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10616_2013_9678_MOESM1_ESM.tif

Estimate of relative abundance of various genes profiled in this study in HP cell line. Time profiles of ΔCT are plotted for a) IgG HC and LC b) Chaperones c) PERK pathway genes and d) Ire1α pathway genes. ΔCT is defined as , Positive values imply abundance greater than β-actin whereas negative values imply lower abundance. Note that β-actin is a moderately abundant gene (TIFF 1084 kb)

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Prashad, K., Mehra, S. Dynamics of unfolded protein response in recombinant CHO cells. Cytotechnology 67, 237–254 (2015). https://doi.org/10.1007/s10616-013-9678-8

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