Functional analysis of cold-inducible cDNA clones in the legume Ammopiptanthus mongolicus

Ammopiptanthus mongolicus is the only evergreen broadleaved shrub endemic species to the Alashan Desert, northwest sand area of China, and can survive at -30℃ or even lower temperature in winter. A modified solid-phase subtraction hybridization technique was developed to isolate and screen cDNAs, whose transcripts increased in cold-treated A. mongolicus seedlings. Sequence analysis of the screened clones indicated that 11 clones had coding regions, with four of them containing complete open reading frame. And nine of the 11 clones shared various degrees of homology with the genes found in the GenBank database and the other two were unidentified sequences. Sequence data further revealed that these accumulated transcripts encode three low molecular weight proteins, a late-embryogenesis protein and two cold acclimation responsive proteins; two photosynthesis-related proteins, photosystem I subunit II precursor (PsaD) and photosystem II oxygen-evolving complex 33kDa subunit (OEC33); a protease inhibitor; an adenosine triphosphatase and a 14-3-3 related protein. Analysis on the function of these proteins indicated that the low molecular weight proteins were associated with water holding ability of cytoplasm; photosynthesis-related proteins participated in the adjustments of photosynthetic apparatus to resist photoinhibition; 14-3-3 related protein could interact with adenosine triphosphatase to enhance ATPase activity and energy metabolism; and protease inhibitor involved in the prevention of unwanted cell death was caused by reactive oxygen species. We suggest that cold acclimation with low light intensity in A. mongolicus is a more complex interaction of low temperature, light, energy and signal than that assumed previously.