DTT-Induced Transcriptome Profiling Reveals Divergent ER Proteostasis and Stress-Adaptation Programs in CHO-S and Apoptosis-Resistant CHO-4BGD Producer Cell Lines

Chinese hamster ovary (CHO) producer cells differ in their capacity to accommodate secretory and endoplasmic reticulum (ER) stress, but how engineered survival backgrounds shape ER-proteostasis responses remains poorly understood. We compared 24-h dithiothreitol (DTT)-induced reductive ER stress responses in two clonal producers: CHO-S-derived HB8 secreting human chorionic gonadotropin and apoptosis-resistant CHO-4BGD-derived DUL secreting a GLP-1–Fc fusion protein with similar specific productivities. Responses were analyzed by strand-specific RNA-seq, Xbp1-splicing RT-PCR, immunoblotting, functional enrichment and curated-module analysis. RNA-seq identified 404 differentially expressed genes in HB8 and 1019 in DUL; all 222 genes shared between two producers changed concordantly. Both producers showed ISR/ATF4/CHOP-associated transcriptional activation and Xbp1 mRNA splicing, whereas changes in total Xbp1 and Hspa5/BiP transcript abundance were limited. BiP and CHOP accumulation was detected at the protein level, while phospho-eIF2α and ATF6 responses were variable. Baseline RNA-seq data comparison identified 1879 differentially expressed genes. HB8 showed higher expression of several classical ER folding/redox factors, whereas DUL showed higher expression of selected ISR-, quality-control- and stress-survival-associated genes. DUL additionally displayed broader vesicle/endocytic and amino-acid/glutathione-related remodeling. Thus, the producers occupy distinct ER-proteostasis states, and DUL mounts a broader, but not uniformly stronger canonical UPR, response to reductive ER stress.