Development of a BiAD sensor for locus-specific detection of cellular histone acetylation dynamics by fluorescence microscopy

Dynamic changes in histone acetylation play crucial roles during cellular differentiation and disease development. Here, we developed a Bimolecular Anchor Detector (BiAD) sensor for the detection of locus-specific changes in histone acetylation in living cells by fluorescence microscopy. For this, we used the BRD9 bromodomain cloned as tandem double-domain (2xBRD9-BD) as reader of histone acetylation. It was integrated it into a dual-color BiAD chassis that was previously described [Kohler et al., 2024, Cell Rep Methods 4(4):100739]. We identified the gene body of TTC34 as potential target for our sensor, because it contains dense histone acetylation and 392 local sequence repeats. Using a binding deficient mutant of 2xBRD9-BD as negative control, we established successful readout of histone acetylation at the TTC34 locus. A single domain reader did not function indicating the requirement for the double reader to enhance affinity and specificity of the chromatin interaction via avidity effects. With this sensor, we could detect dynamic changes of histone acetylation at the TTC34 locus after treatment of cells with the histone deacetylase inhibitor Trichostatin A for 6 hours indicating the applicability of this sensor for single-cell epigenome studies.