Accurate control of gene transcription is necessary for most cellular processes and dysregulation of transcription underlies development and progression of many human diseases. Gene transcription is regulated by a complex interplay of sequence-specific transcription factors, coactivator complexes, and chromatin landscape. Chromatin readers are regulatory factors which translate modifications on histone tails into specific transcriptional output. Most chromatin readers are subunits of large protein complexes which remodel or modify chromatin. One of the exceptions is a bromodomain and extra-terminal domain (BET) protein family. BET proteins are characterized by the presence of two bromodomain which allows them to recognize acetylated lysine residues and an extraterminal (ET) domain which was proposed to support protein-protein interactions. BET family members are conserved across eukaryotes from yeast to humans and are required for maintaining global transcriptional programs in both healthy and diseased cells. Consequently, BET proteins are a promising drug target for many human pathologies. Despite their essential function in transcriptional regulation the specific roles of BET proteins are not well understood. BET proteins have a modular architecture which includes bromodomains, ET domain and several other highly conserved domains of mostly unknown biological significance. Our central hypothesis, supported by preliminary data, assumes that BET protein domains other than bromodomains are important for the roles of BET proteins in transcriptional regulation. We propose to employ the yeast model system for a comprehensive investigation into the functions of all BET domains conserved between yeast and human cells. Building on our prior work, we will focus on the mechanisms which direct BET protein association with chromatin and participation of BET proteins in transcription initiation. We speculate that BET proteins provide an interaction hub for recruitment of coactivator complexes during transcription initiation, and we expect that specific domains of BET factors contribute to these activities. The results of the project will significantly expand the models of transcriptional regulation in both yeast and human cells. Characterization of the functions of individual BET protein domains may also lead to identification of promising drug targets to modulate aberrant transcription and offer an alternative to non-selective inhibition of BET bromodomains.