The BET family inhibitor JQ1 specifically targets FSH in Drosophila. (A) Conserved protein domain arrangement between BET family proteins. The Drosophila BET family is exclusively represented by the long and short isoform of FSH. BD1: Bromodomain 1; BD2: Bromodomain 2; CTM: C-terminal motive; ET: Extraterminal domain. (B) Assesment of BET inhibitor selectivity using a phylogenetic analysis of Drosophila (blue) and human (black) bromodomains. BRD and FSH bromodomains form the BET clade highligted in gray. Notecolors of human proteins indicate averaged temperature shifts upon binding of 10 µM JQ1 measured by differential scanning fluorimerty . (C) Protein sequence alignment of human and Drosophila BET bromodomains. Boxed residues reside in α helices as determined by structural analysis . The high degree of sequence conservation suggests that FSH bromodomains shape into the typical left-handed bundle of four α helices (αZ, αA, αB, αC) connected by loop regions. (D) Histone peptide pull-down assays using a purified, 6xHis-tagged BD1-BD2 fragment from FSH. 10 µM JQ1 effectively eliminates the interaction between acetlyted H4 (K5, K8, K12, K16) or H3 (K9, K14) peptides and the di-bromodomain fragment. (E) Similar histone peptide pull-down assay using nuclear extract from HEK293 cells transfected with V5-tagged FSH-S.