Journal
CURRENT RESEARCH IN BIOTECHNOLOGY
Volume 1, Issue -, Pages 49-57Publisher
ELSEVIER
DOI: 10.1016/j.crbiot.2019.09.001
Keywords
Chromatin immunoprecipitation (ChIP); CHO cell line selection; Nuclear proteomics; Transcriptional regulation
Categories
Funding
- U.S. National Science Foundation [NSF CBET-096782]
- Fulbright Global Scholar Award
- European Union [642663]
- Science Foundation Ireland Infrastructure Award [16/RI/3701]
- Novo Nordisk Foundation [NNF10CC1016517]
- U.S. National Institutes of Health NIGMS [R35 GM119850]
- Science Foundation Ireland (SFI) [16/RI/3701] Funding Source: Science Foundation Ireland (SFI)
- Marie Curie Actions (MSCA) [642663] Funding Source: Marie Curie Actions (MSCA)
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Most therapeutic monoclonal antibodies in biopharmaceutical processes are produced in Chinese hamster ovary (CHO) cells. Technological advances have rendered the selection procedure for higher producers a robust protocol. However, information on molecular mechanisms that impart the property of hyper-productivity in the final selected clones is currently lacking. In this study, an IgG-producing industrial cell line and its methotrexate (MTX)-amplified progeny cell line were analyzed using transcriptomic, proteomic, phosphoproteomic, and chromatin immunoprecipitation (ChIP) techniques. Computational prediction of transcription factor binding to the transgene cytomegalovirus (CMV) promoter by the Transcription Element Search System and upstream regulator analysis of the differential transcriptomic data suggested increased in vivo CMV promoter-cAMP response element binding protein (CREB1) interaction in the higher producing cell line. Differential nuclear proteomic analysis detected 1.3-fold less CREB1 in the nucleus of the high productivity cell line compared with the parental cell line. However, the differential abundance of multiple CREB1 phosphopeptides suggested an increase in CREB1 activity in the higher producing cell line, which was confirmed by increased association of the CMV promotor with CREB1 in the high producer cell line. Thus, we show here that the nuclear proteome and phosphoproteome have an important role in regulating final productivity of recombinant proteins from CHO cells, and that CREB1 may play a role in transcriptional enhancement. Moreover, CREB1 phosphosites may be potential targets for cell engineering for increased productivity.
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