Transgenic overexpression of the transcription factor Alfin1 enhances expression of the endogenous MsPRP2 gene in alfalfa and improves salinity tolerance of the plants

Alfin1 cDNA encodes a putative transcription factor associated with NaCl tolerance in alfalfa (Medicago sativa L.). The recombinant protein binds DNA in a sequence-specific manner, including promoter fragments of the NaCl-inducible gene MsPRP2. Alfin1 function was tested in transgenic alfalfa under the control of the 35S promoter in the sense and antisense orientations with the endogenous MsPRP2 as a reporter gene. Calli overexpressing Alfin1 were more resistant to growth inhibition by 171 mM NaCl than vector-transformed controls, whereas calli expressing Alfin1 in the antisense orientation were more sensitive to NaCl inhibition. Transgenic plants overexpressing Alfin1 in the sense orientation grew well. In contrast, the antisense transgenic plants grew poorly in soil, demonstrating that Alfin1 expression is essential for normal plant development. Transgenic calli and plant roots overexpressing Alfin1 showed enhanced levels of endogenous MsPRP2 mRNA accumulation. However, MsPRP2 mRNA accumulation was also regulated in a tissue-specific manner, as shown in leaves of transgenic plants overexpressing Alfin1. These results suggest that Alfin1 acts as a transcriptional regulator in plants and regulates MsPRP2 expression in alfalfa. Alfin1 overexpressing transgenic plants showed salinity tolerance comparable to one of our NaCl-tolerant plants, indicating that Alfin1 also functions in gene regulation in NaCl tolerance.