The putative cofactor TIF1α is a protein kinase that is hyperphosphorylated upon interaction with liganded nuclear receptors

Abstract

Ligand-induced gene activation by nuclear receptors (NRs) is a complex process requiring dissociation of corepressors and recruitment of coactivators. The putative transcriptional intermediary factor TIF1α has been previously characterized as a nuclear protein that interacts directly with the AF-2 ligand-dependent activating domain present in the ligand- binding domain of numerous steroid and nonsteroid receptors, including the estrogen (ERα) and retinoid X (RXRα) receptors. We report here that TIF1α is both a phosphoprotein and a protein kinase. TIF1α coexpressed in COS-1 cells with RXRα or ERα is phosphorylated and becomes hyperphosphorylated upon ligand treatment. This hyperphosphorylation requires the binding of TIF1α to transcriptionally active NRs since it is prevented by mutations either in the core (α-helix 12 of the ligand-binding domain) of the AF-2 activating domains of RXRα and ERα or in the NR box of TIF1α that are known to prevent TIFlα-NR interactions. Thus, TIF1α is a phosphoprotein that undergoes ligand-dependent hyperphosphorylation as a consequence of nuclear receptor binding. We further show that purified recombinant TIF1α possesses intrinsic kinase activity and that, in addition to autophosphorylation, TIFlα selectively phosphorylates the transcription factors TFIIEα, TAF(II)28, and TAF(II)55 in vitro. These latter results raise the possibility that TIF1α may act, at least in part, by phosphorylating and modifying the activity of components of the transcriptional machinery.