In mammalian cells, up to six serines and threonines in histone H1 are phosphorylated in vivo in a cell cycle dependent manner that has long been linked with chromatin condensation. Growth-associated H1 kinases, now known as cyclin-dependent kinases (CDKs), are thought to be the enzymes responsible for this process. This paper describes the phosphorylation of histone H1 by four different purified CDKs. The four CDKs phosphorylate only the cell cycle specific phosphorylation sites of H1, indicating that they belong to the kinase class responsible for growth-related H1 phosphorylation in vivo. All four CDKs phosphorylate all of the interphase and mitotic-specific H1 sites. In addition to the (S/T)PXK consensus phosphorylation sites, these four CDKs also phosphorylate a mitotic-specific in vivo H1 phosphorylation site that lacks this sequence. There is no site selectivity among the growth-related phosphorylation sites by any of the four CDKs because all four CDKs phosphorylate all relevant sites. The results imply that the cell cycle dependent H1 phosphorylations observed in vivo must involve differential accessibility of H1 sites at different stages of the cell cycle.