In Tetrahymena, histone H1 phosphorylation can regulate transcription and mimics loss of H1 from chromatin. We investigated the mechanism by which H1 phosphorylation affects transcription. Tetrahymena strains were created containing mutations in H1 that mimicked the charge of the phosphorylated region without mimicking the structure or increased hydrophilicity of the phosphorylated residues. Whenever the charge resembled that of the phosphorylated state, the induced expression of the CyP1 gene was greatly inhibited. Whenever the charge was similar to that of the dephosphorylated state, the CyP1 gene was induced normally. These results argue strongly that phosphorylation of H1 acts by changing the overall charge of a small domain, not by phosphate recognition or by creating a site-specific charge.