Document Detail


Quantitative analysis of transcription and translation in gene amplified Chinese hamster ovary cells on the basis of a kinetic model.
MedLine Citation:
PMID:  22359058     Owner:  NLM     Status:  PubMed-not-MEDLINE    
Abstract/OtherAbstract:
The elevation of expression levels for secreted glycoproteins by gene amplification in mammalian cells shows a saturation behavior at high levels of gene amplification. At high expression levels a drop in the secretion efficiency for the recombinant protein occurs (Schröder and Friedl, 1997), coinciding with the appearance of misfolded protein in the cell. In this communication we investigated whether additional limitations exist at the levels of transcription and translation. Four Chinese hamster ovary (CHO) cell lines expressing different amounts of human antithrombin III (ATIII) were used as a model system. A tenfold increase in the ATIII cDNA copy number from the lowest to the highest producing cell line coincided with a 38-fold increase in ATIII mRNA levels, and an 80-fold increase in the amount of intracellular ATIII levels. The data was analyzed using a simple kinetic model. The following conclusions were derived: I. The transcriptional activity for the recombinant protein is not saturated. II. Translation itself is not saturated either, but may be downregulated as secretion efficiency drops. III. Two explanations for the previously reported drop in secretion efficiency for the recombinant protein with increasing expression level are possible: A. Protein degradation is an alternative fate for translated ATIII and the fraction of ATIII degraded after translation increases as expression level is increased. B. Translation is downregulated as the secretory apparatus becomes exhausted to maintain cell viability.
Authors:
M Schröder; C Körner; P Friedl
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Cytotechnology     Volume:  29     ISSN:  0920-9069     ISO Abbreviation:  Cytotechnology     Publication Date:  1999 Mar 
Date Detail:
Created Date:  2012-02-23     Completed Date:  2012-10-02     Revised Date:  2013-03-07    
Medline Journal Info:
Nlm Unique ID:  8807027     Medline TA:  Cytotechnology     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  93-102     Citation Subset:  -    
Affiliation:
Institut für Biochemie, Technische Hochschule Darmstadt, Petersenstraße 22, D-64287, Darmstadt, Germany E-mail, meschrod@umich.edu.
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