Multiple receptor interaction domains of GRIP1 function in synergy.

Nuclear hormone receptors are exerting their effect on transcription by interacting with basal factors of the transcription machinery and/or by recruiting intermediary factors, such as the mouse protein GRIP1. GRIP1 is one of the recently identified coactivators for nuclear hormone receptors. Upon interaction with the hormone-binding domain of the receptors, GRIP1 increases their transcriptional activity. Here we show that GRIP1 contains at least two receptor-interacting regions using the hormone-binding domain of several receptors as bait in the yeast two-hybrid assay. GRIP1 interacts in a hormone-dependent manner with the C-termini of nuclear hormone receptors such as GRalpha, TRalpha, TRbeta, RARalpha and RXRalpha but not with v-ErbA. GRIP1 contains several LXXLL motifs which were shown to be required for receptor interaction. A protein fragment containing all of the three LXXLL motifs, but having the activation domain deleted, is able to repress the transcriptional activity of human TRbeta, whereas a region harbouring only one LXXLL motif fails to do so. A protein fragment with two LXXLL motifs exhibits an intermediate modulation of the TRbeta transactivation. While one motif seems to be sufficient for receptor interaction, more than one motif is needed for functional interference.