where all three are evicted is not observed, indicating that these eviction events are not independent (Brown et al. 2013; Brown and Boeger 2014). In another study, DNA methyltransferase was used to footprint nucleosomes in individual cells, with bisulfite sequencing (effectively a single-molecule technique) being used to assess nucleosome occupancy across a 500- to 600-bp region. Using this method, analysis of activated PHO5 and two other promoters reveals activated configurations in which the promoter nucleosomes are not ejected, but their position is shifted in a coordinated manner (Small et al. 2014). They also show that the frequency of different configurations changes upon transcriptional activation. Detailed single-cell studies of nucleosome configurations can increase our understanding of how nucleosome positioning impacts genomic processes. A promising recent advance is the development of single-cell ATAC-seq, which provides single-cell analysis of nucleosome depletion in metazoans and reveals subpopulations of cell states within a given cell type (Buenrostro et al. 2015; Cusanovich et al. 2015), although as ATAC-seq primarily provides information on nucleosome depletion, it does not address the questions of nucleosome configurational diversity raised above.
