Recombinant Chinese hamster ovary (CHO) cells selected for high productivity are capable of secreting immunoglobulin G (IgG) molecules at a level that rivals plasma cells in vivo. Following butyrate treatment at 33 °C, further increases in productivity are observed. To better understand the mechanisms by which this increased productivity is incurred, the transcriptional response of an antibody-producing cell line undergoing these treatments was investigated using oligo-DNA microarrays. Using distance calculations, more than 900 genes were identified as kinetically differentially expressed between the butyrate-treated 33 °C culture and the untreated culture. Furthermore, transcript levels of the heavy and light chain IgG genes increased following treatment. Using stable isotope labeling (SILAC), the secretion rate of IgG was investigated by tracking the decay of the isotope label upon switching to unlabeled medium. Both treated and untreated cultures exhibited very similar IgG secretion kinetics. In contrast, the intracellular IgG content was found to be elevated following treatment. This result suggests that increased productivity under treatment is attributable to elevated cellular secretory capacity, rather than shorter holding times in the secretory pathway. This hypothesis is further supported by the results of gene set enrichment analysis (GSEA), which revealed that elements of the secretory pathway, including Golgi apparatus, cytoskeleton protein binding and small GTPase-mediated signal transduction are enriched and thus may play a role in the increased recombinant protein production observed under butyrate treatment at 33 °C.
Bibliographical noteFunding Information:
AK was supported by the NIH Biotechnology Training Grant ( GM08347 ). The authors would like to acknowledge Dr. Peter Morin Nissom and Song Hui Chuah (BTI, A*STAR, Singapore) for assistance in the construction of the CHO Affymetrix array. We also thank Dr. Lorraine B. Anderson, Dr. LeeAnn Higgins and Dr. Bruce Witthunh of the Center for Mass Spectrometry and Proteomics (University of Minnesota, USA) for their assistance with mass spectrometry. The authors would also like to acknowledge Professor Timothy J. Griffin and Dr. Karthik Jayapal for insightful discussions. Bioinformatics support was provided by the University of Minnesota Supercomputing Institute.
Copyright 2010 Elsevier B.V., All rights reserved.
- CHO cells
- DNA microarrays
- Gene set enrichment
- Mammalian cell culture
- Secretion kinetics
- Sodium butyrate