The Early Growth Response Genes (EGR-1 to AT133/EGR-4) encode a family of proteins that are composed of three homologous consecutive zinc fingers of the Cys2-His2 type and different flanking sequence. Upon growth stimulation of resting cells the four EGR-genes are simultaneously transcribed. We have analyzed the expression of the four EGR-proteins in Jurkat T cells and show by Western blot analysis that the four EGR-proteins are coordinately induced upon treatment with a combination of PHA and PMA. As the individual proteins are reported to bind to identical target sequences, we have analyzed the DNA-binding of the native proteins. Using nuclear extract in which we have demonstrated expression of all four EGR-proteins, only EGR-1, but no other member of this protein family is found to bind to the EGR-consensus site (GCG GGG GCG). In addition, DNA-binding of both native EGR-1 and of recombinant EGR-1 and AT133/EGR-4 proteins expressed in insect cells was analyzed. This comparison revealed distinct binding properties of recombinant EGR-1 and AT133/EGR-4 to oligonucleotides that include the EGR-consensus sites. The distinct binding affinities suggest that in vivo EGR-proteins bind to different target sequences and that each EGR-protein regulates distinct target genes. This is underlined by demonstrating that EGR-1 but not AT133/EGR-4 binds to a related G-rich promoter element with the sequence GGG GTG GGG. This G-rich sequence serves as an overlapping binding site for the two zinc finger proteins EGR-1 and Sp1. As similar overlapping binding sites for EGR-1 and Sp1 have been identified in several human and mouse gene promoters, we raise the question whether the Sp1 binding sites described in a large number of eukaryotic gene promoters also represent binding sites for EGR-1.