Multiplexed Digital mRNA Expression Analysis Profiles System-Wide Changes in mRNA Abundance and Responsiveness of UPR-Specific Gene Expression Changes During Batch Culture of Recombinant Chinese Hamster Ovary Cells.

Abstract

The unfolded protein response (UPR) signaling pathway is viewed as critical for setting the effectiveness of recombinant protein expression in CHO cells. In this study, Nanostring nCounter technology is used to study expression of a group of genes associated with cellular processes linked to UPR activation under ER stress and the changing environment of a batch culture. Time course induction of ER stress, using tunicamycin (TM), shows a group of genes such as Chop, Trb3, Sqstm1, Grp78, and Herpud1 respond rapidly to TM inhibition of N-glycosylation, while others such as Atf5, Odz4, and Birc5 exhibits a delayed response. In batch culture, expression of "classical" UPR markers only increases when cells enter decline phase. In addition to providing a detailed analysis of the expression of process-relevant UPR markers during batch culture and in response to imposed chemical stress, we also highlighted six genes (Herpud1, Odz4, Sqstm1, Trb3, Syvn1, and Birc5) associated with the perception of ER stress responses in recombinant CHO cells. Herpud1 (involved in ER-associated degradation) exhibits a rapid (primary) response to stress and its relationship (and that of the other five genes) to the overall cellular UPR may identify novel targets to modulate recombinant protein production in CHO cells.