Cell signaling that culminates in posttranslational modifications directs protein activity. Here we report how multiple Ca2+-dependent phosphorylation sites within the transcription activator Ets-1 act additively to produce graded DNA binding affinity. Nuclear magnetic resonance spectroscopic analyses show that phosphorylation shifts Ets-1 from a dynamic conformation poised to bind DNA to a well-folded inhibited state. These phosphates lie in an unstructured flexible region that functions as the allosteric effector of autoinhibition. Variable phosphorylation thus serves as a ``rheostat'' for cell signaling to fine-tune transcription at the level of DNA binding.