Phosphorylation of linker histones by cAMP-dependent protein kinase in mitotic micronuclei of Tetrahymena

Abstract

Linker histones (LHs) in transcriptionally inretive, mitotically dividing micronuclei of Tetrahymena thermophila, α, β, γ and δ, are highly phosphorylated in vivo. Analysis of the derived sequences of these LHs suggests that none of these polypeptides contain sites of phosphorylation by p34^cdc2, the kinase thought to play an essential role governing the entry of all cells into mitosis. Suprisingly α, β, γ and δ each contain sites for phosphorylation by cyclic AMP-dependent kinase (PKA). p34^cdc2 kinase phosphorylases H1 in vitro but fails to phosphorylate α, β, γ and δ. Conversely, PKA phosphorylates each of the micronuclear LHs but is unable to phosphorylate macronuclear H1. Micronuclear LHs labeled in vivo with [³²P]phosphate were purified by reverse phase HPLC. Phosphoamino acid analysis showed that all four micronuclear LHs are phosphorylated exclusively on serine residues in vitro. Cyanogen bromide mapping of α, β, γ and δ labeled in vivo or in vitro by PKA indicates that each LH is phosphorylated only on peptides that contain either optimum (RR/KXS) or less optimum (RXXS) PKA sequences. This study suggests that PKA or a PKA-like activity(ies), but not p34^cdc2 kinase, is(are) responsible for the in vivo phosphorylation of LHs in the mitotic micronucleus of Tetrahymena. We suggest that, at least in Tetrahymena, PKA-driven phosphorylation or dephosphorylation plays a significant role in the control of mitotic processes such as chromosome condensation.