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A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.).

Bertrand, A., Bipfubusa, M., Castonguay, Y., Rocher, S., Szopinska-Morawska, A., Papadopoulos, Y. et Renaut, J. 2016. A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.). BMC Plant Biology.


Improvement of freezing tolerance of red clover (Trifolium pratense L.) would increase its persistence under harsh winter conditions. In this study, we assessed the freezing tolerance and compared the proteome composition of non-acclimated and cold-acclimated plants of TF0 populations and of populations TF3 and TF4 obtained respectively after three and four cycles of phenotypic recurrent selection for superior freezing tolerance in two initial backgrounds of red clover: cultivars Endure (E-TF0) and Christie (C-TF0). Plants were hardened to natural variation in air and soil temperatures throughout fall and winter in an unheated greenhouse. Freezing tolerance expressed as the lethal temperature for 50% of the plants (LT50) increased markedly from approximately -2 to -16 °C following cold hardening. Recurrent selection allowed a significant 2 to 3°C increase of the LT50 after four cycles of recurrent selection. Difference gel electrophoresis (DIGE) based on fluorochromes labelling and separation by two-dimensional electrophoresis was used to study variations in protein abundance. Principal component analysis based on DIGE revealed that the largest variability in the protein data set was attributable to the cold acclimation treatment. The two genetic backgrounds (E and C) which were clustered in the non-acclimated state were differentiated after cold acclimation. Proteins that vary in abundance after cold acclimation, between genetic backgrounds or in response to selection were identified by mass spectrometry. A marked increase in vegetative storage proteins (VSP) and dehydrins was among the most striking changes in proteome composition of cold acclimated crowns of red clovers. A subset of proteins varied in abundance in response to selection including a KS-dehydrin that increased in abundance in TF3 and TF4 populations as compared to TF0 in the Endure background. Our observations suggest that variation in freezing tolerance in the two genetic backgrounds are associated with both common and specific changes in cold-acclimated protein complements.

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