The cold‐shock response in cultured mammalian cells: Harnessing the response for the improvement of recombinant protein production

Abstract

There are a growing number of reports on the sub-physiological temperature culturing (<37 degrees C) of mammalian cells for increased recombinant protein yield, although the effect is variable between cell lines, expression systems, and the product of interest. What is becoming clear is that exposing mammalian cells to sub-physiological temperatures invokes a coordinated cellular response involving modulation of the cell cycle, metabolism, transcription, translation, and the cell cytoskeleton. Opportunities currently exist for further enhancement of the cold-shock effect on recombinant protein production in mammalian cells through advancements in our understanding of the mechanisms involved in the cold-shock response.

Proliferation control strategies to improve productivity and survival during CHO based production culture

“…It has been proven that it is an active response and cells synthesize specific cold inducible proteins (Kondo et al 1992;Holland et al 1993;Jones and Inouye 1994;Derry et al 1995;Nishiyama et al 1997a, b;Kaufmann et al 1999;Danno et al 2000;Sonna et al 2002;Dresios et al 2005;Yoon et al 2006;Baik et al 2006;Al-Fageeh et al 2006). A change in the phosphorylation patterns at the tyrosine residues of two proteins (~180 and 80 kDa) was also observed following reduction of the culture temperature providing further proof that there is an active signalling response to mild hypothermia in CHO (Kaufmann et al 1999).…”

“…The quality of recombinant products, with regard to isoform pattern, sialic acid content, and in vivo biological activity, is maintained at reduced temperatures (Yoon et al 2003b;Clark et al 2004;Bollati-Fogolin et al 2005). The mechanism whereby cells at lower temperatures improve productivity is still poorly understood although recent studies have demonstrated that reduced culture temperature invokes a coordi- nated response involving the cell cycle, transcription and translational machinery, and the arrangement of the cytoskeleton (Chuppa et al 1997;Moore et al 1997;Ohsuye 1998, 1999;Jorjani and Ozturk 1999;Hendrick et al 2001;Yoon et al 2003b;Fox et al 2004Fox et al , 2005Baik et al 2006;Al-Fageeh et al 2006). Low culture temperature results in reduced metabolism (glucose/medium consumption, oxygen uptake, lactate production and ammonia production) and shear sensitivity which leads to delayed initiation of apoptosis and helps in extending the duration of the stationary/production phase (Moore et al 1997;Chuppa et al 1997;Ohsuye 1998, 1999;Jorjani and Ozturk 1999;Hendrick et al 2001;Yoon et al 2003a, b;Trummer et al 2006;Wong et al 2006).…”

“…Although global protein translation is reduced at lower temperatures in mammalian cells (Wassmann et al 1998;Phadtare et al 1999), more than 20 cold inducible proteins have been identified, including cold-inducible RNA binding protein (CIRP, also known as hnRNP A18), RNA binding motif protein 3 (RBM3), protein disulphide isomerase (PDI), NSR1, vimentin, NDK B, ERp57, gi-27754065, phosphoglycerate kinase, heat shock cognate 71 kDa protein, HSP90-b and EF2, (Nishiyama et al 1997a, b;Sheikh et al 1997;Kondo et al 1992;Danno et al 2000;Sonna et al 2002;Al-Fageeh et al 2006;Baik et al 2006). In this laboratory global proteomic analysis of cells during exponential phase growth at 37°C and low temperature induced stationary phase at 31°C has resulted in the identification of 201 differentially expressed proteins out of 2,852 detected, of which 118 were upregulated (Kumar et al in preparation).…”

“…Therefore although the effects of low culture temperature on the cellular productivity of recombinant proteins are variable and depend on the temperature, cell line, clone, expression system and/or protein, the effect of reducing cell growth is a universal phenomenon Ohsuye 1998, 1999;Kaufmann et al 1999;Ryll et al 2000;Schatz et al 2003;Yoon et al 2003aYoon et al , b, 2004Yoon et al , 2006Fox et al 2004;Fogolin et al 2004, Bollati-Fogolin et al 2005Trummer et al 2006;Al-Fageeh et al 2006). …”

Expression of recombinant Atlantic salmon serum C-type lectin in Drosophila melanogaster Schneider 2 cells

“…Other possibilities, including manipulation of the temperature for example, could also enhance production. Low temperature appears to hold promise for enhancing protein expression yields in mammalian cells (Al-Fageeh et al 2006;Wulhfard et al 2008). Nonetheless, the production of rSSL as a secreted product in a eukaryotic expression system might remain difficult as a result of its function.…”