The sfr mutations, which result in sensitivity to freezing after cold acclimation, define genes that are required for freezing tolerance. We tested plants homozygous for mutations sfr2 to sfr7 for cold-induced gene expression and found that sfr 6 plants were deficient in cold-inducible expression of the genes KIN1, COR15a, and LTI78, which all contain the C repeat/dehydration-responsive element (CRT/DRE) motif in their promoters. Similarly, sfr 6 plants failed to induce KIN1 normally in response to either osmotic stress or the application of abscisic acid. In contrast, cold-inducible expression of genes CBF1, CBF2, CBF3, and ATP5CS1, which lack the CRT/DRE motif, was not affected. The freezing-sensitive phenotype that defines sfr 6 also was found to be tightly linked to the gene expression phenotype. To determine whether the failure of cold induction of CRT/DRE-containing genes in sfr 6 was due to altered low-temperature calcium signaling, cold-induced cytosolic-free calcium ([Ca2+]cyt) elevations were investigated in the sfr 6 mutant, but these were found to be indistinguishable from those of the wild type. We discuss the possibilities that CRT/DRE binding proteins (such as CBF1) require activation to play a role in transcription and that the SFR6 protein is a vital component of their activation.