Activator-binding domains are required but insufficient for Med15 recruitment to UAS, revealing a critical role for its disordered C-terminus in directing genome targeting

The Mediator complex is a conserved transcriptional coactivator that bridges transcription factors (TFs) with the core transcriptional machinery. In budding yeast, TFs recruit the Mediator to upstream activation sequences (UASs) by interacting with three activator-binding domains (ABDs) within the Med15 tail subunit. However, these ABD-TF interactions exhibit relatively low affinity, raising the question of whether they fully account for Med15 recruitment in vivo. Here, we address this question by combining genome-wide profiling with systematic mutational analysis. We demonstrate that while the three ABDs are collectively required for Med15 recruitment, they are not individually required and are not sufficient to specify the genomic binding pattern of Med15. Instead, removal of an intrinsically disordered C-terminal redistributes Med15 occupancy across UASs. Notably, C-terminal truncation alters Med15 binding even at TF-depleted regions, and these effects are recapitulated by deleting Med2 or Med3, which further stabilize TF-dependent recruitment. Together, these findings support a dual mechanism of Med15 UAS recruitment, whereby TF-ABD recruitment is directed and stabilized by C-terminal mediated interactions.