Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. To understand how MITF regulates transcription, we used tandem affinity purification and mass spectrometry to define a comprehensive MITF interactome identifying novel cofactors involved in transcription, DNA replication and repair, and chromatin organisation. We show that MITF interacts with a PBAF chromatin remodelling complex comprising BRG1 and CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. Combinations of MITF, SOX10, TFAP2A, and YY1 bind between two BRG1-occupied nucleosomes thus defining both a signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of the regulatory elements they occupy. BRG1 also regulates the dynamics of MITF genomic occupancy. MITF-BRG1 interplay thus plays an essential role in transcription regulation in melanoma.DOI: http://dx.doi.org/10.7554/eLife.06857.001

We have cloned cDNAs encoding three novel TAFIIs [TATA‐binding protein (TBP)‐associated factors] from the human (h) HeLa cell TFIID complexes hTAFII28, hTAFII20 and hTAFII18. hTAFII28 is a core hTAFII present in both of the previously described hTFIID species which either lack or contain hTAFII30 (hTFIID alpha and hTFIID beta respectively), and is the homologue of Drosophila (d)TAFII30 beta. hTAFII18 is a novel hTAFII which shows homology to the N‐terminal region of the yeast TAFIISPT3, but has no known Drosophila counterpart. In contrast to hTAFII28, hTAFII18 is a TFIID beta‐specific hTAFII. hTAFII20 is the homologue of p22, an alternatively spliced form of dTAFII30 alpha (p32). Using a combination of protein affinity chromatography and cotransfection and immunoprecipitation assays, we have identified a series of in vitro and intracellular interactions among the novel hTAFIIs and between the novel hTAFIIs and hTAFII30 or TBP. We show that hTAFII28 interacts with hTAFII18 both in vitro and intracellularly; in contrast to its Drosophila homologue, hTAFII28 also interacts directly with TBP. Deletion analysis indicates that TBP and hTAFII18 bind to distinct domains of hTAFII28. hTAFII18 also interacts with TBP, but it interacts more strongly with hTAFII28 and hTAFII30. The binding of hTAFII28 and hTAFII30 requires distinct domains of hTAFII18. As observed with the homologous Drosophila proteins, hTAFII20 interacts directly with TBP; however, additional interactions between hTAFII20 and hTAFII28 or hTAFII30 were detected. These results reveal differences not only in subunit composition, but also in the organization of dTFIID and hTFIID complexes.

Dosage compensation in flies involves doubling the transcription of genes on the single male X chromosome to match the combined expression level of the two female X chromosomes. Crucial for this activation is the acetylation of histone H4 by the histone acetyltransferase (HAT) MOF. In male cells, MOF resides in a complex (dosage compensation complex, DCC) with MSL proteins and noncoding roX RNA. Previous studies suggested that MOF's localization to the X chromosome was largely RNA-mediated. We now found that contact of the MOF chromo-related domain with roX RNA plays only a minor role in correct targeting to the X chromosome in vivo. Instead, a strong, direct interaction between a conserved MSL1 domain and a zinc finger within MOF's HAT domain is crucial. The functional consequences of this interaction were studied in vitro. Simultaneous contact of MOF with MSL1 and MSL3 led to its recruitment to chromatin, a dramatic stimulation of HAT activity and to improved substrate specificity. Activation of MOF's HAT activity upon integration into the DCC may serve to restrict the critical histone modification to the male X chromosome.