Histone occupancy in vivo at the 601 nucleosome binding element is determined by transcriptional history.

We report in vivo analysis of histone and RNA polymerase II (pol II) occupancy at the 601 element, which functions as a strong in vitro nucleosome-positioning element and transcriptional pause site. Surprisingly, nucleosomes were not strongly positioned over the 601 element inserted either within a yeast chromosomal open reading frame (ORF) (GAL1-YLR454W) or in an intergenic region. In fact 601 within GAL1-YLR454W was actually depleted of histones relative to flanking sequences and did not cause pol II pausing. Upstream of an inserted 601 element within GAL1-YLR454W, a positioned nucleosome was formed whose location depended on transcriptional history; it shifted after a round of activation and repression. Transcriptional activation caused histone eviction throughout the GAL1-YLR454W ORF, except at 601, where there was no loss and some net histone deposition. In contrast, a second round of activation after glucose shutoff caused histone eviction both at 601 and elsewhere in the ORF. We conclude that the intrinsic high-affinity histone-DNA interactions at 601 do not necessarily play a dominant role in establishing nucleosomes or pol II pause sites within a coding region in vivo and that transcriptional history can have an important influence on histone occupancy flanking this sequence.