Expression of freeze-responsive proteins, Fr10 and Li16, from freeze-tolerant frogs enhances freezing survival of BmN insect cells
To date, two novel freeze-responsive proteins, Fr10 and Li16, have been discovered in the wood frog, Rana sylvatica, and likely support freezing survival. Although previous studies have established tissue distribution of each protein, there have been no studies that explore their functional consequences in intolerant cells. To assess the ability of Fr10 and Li16 to confer freeze tolerance, we transfected each protein into a freeze-intolerant silkworm cell line (BmN). Selected controls were the transfection of an unrelated protein (CAT) and a no-transfection sample. Li16 and Fr10 showed 1.8±0.1-and1. 7±0.2-fold, respectively, greater survival after freezing at 6°C for 1 h than did transfection controls. To investigate how these novel proteins protect cells from freezing damage, protein structures were predicted from primary amino acid sequences. Analysis of the structures indicated that Fr10 is a secreted protein and may be a new type IV antifreeze protein, whereas Li16 may have intracellular membrane associated functions. This study shows that freezing protection can be provided to intolerant cells by the overexpression of transfected Li16 and Fr10 frog proteins. Results from this study will provide new insights into adapting intolerant cells for medical organ cryoprotection using a natural vertebrate model of tolerance.
|Journal||The FASEB journal : official publication of the Federation of American Societies for Experimental Biology|
Biggar, K.K, Kotani, E. (Eiji), Furusawa, T. (Toshiharu), & Storey, K. (2013). Expression of freeze-responsive proteins, Fr10 and Li16, from freeze-tolerant frogs enhances freezing survival of BmN insect cells. The FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 27(8), 3376–3383. doi:10.1096/fj.13-230573