cDNA Encoding Putative Zinc Finger Motifs from Salt-Tolerant Alfalfa (Medicago sativa L.) Cells

I have selected salt-tolerant cell lines from salt-sensitive cell cultures to identify genes that can contribute to longt e m increased salt tolerance in alfalfa (Medicago sativa L.). Comparisons of differential gene expression in response to salt within the same genetic background allowed me to identify gene products that showed salt-induced expression in those cells with acquired salt tolerance (Winicov et al., 1989; Winicov and Button, 1991). Salt-tolerant cells, which had been selected with a single-step selection protocol, showed both constitutive and salt-inducible increases in gene expression. Most of these changes were different from those seen in the salt-sensitive parent cells during short-term salt stress. Although very little is known of the functions of proteins induced in conditions of either salt stress or salt tolerance, we hypothesized (Winicov, 1990; Winicov and Button, 1991) that the large-scale enhanced expression of genes in the salt-tolerant cells reflected coordinate changes in regulation of genes for physiological systems that may contribute to the increased tolerance of the selected cells. Specific genes that show enhanced expression in the salttolerant cells were isolated as cDNA clones from a library constructed from poly(A)+ RNA isolated from salt-tolerant cells grown in the presence of salt. In this paper, I report the characterization and sequencing of one cDNA clone isolated by this procedure. Although the function of the translated polypeptide encoded by the Alfin-1 cDNA remains to be identified, it contains two amino acid sequences characteristic of nucleic acid-binding proteins important in gene regulation. Starting from position 184, there is a region of highly concentrated negative charge enriched in Glu and Asp with 12 of the 15 amino acids negatively charged. A similar highly negatively charged region has been shown to be necessary for CAL4 activation of transcription in yeast and is located near the DNA-binding zinc finger in GAL4 (Ma et al., 1988). The negatively charged region is immediately followed by a sequence that is characteristic of metal-binding proteins, commonly referred to as the zinc finger sequence, with one potential Cysd zinc finger and another possible His/Cys3 structure (reviewed by Berg, 1990). Zinc finger domains in proteins have been shown to interact with nucleic acids in gene regulation for both prokaryotic