FRET analysis of unwrapping of nucleosomal DNA containing the sequence characteristic to the +1 nucleosome

Abstract

Sequence-dependent mechanical properties of the DNA could play essential roles in nuclear processes by affecting the histone-DNA interactions. Previously, we found that the DNA entry site of the first nucleosomes from the transcription start site (+ 1 nucleosome) in budding yeast enriches AA/TT steps but not in the exit site, indicating the association with the gene activation. Because AA/TT is a rigid dinucleotide step, we considered that AA/TT causes DNA unwrapping. However, the MNase-Seq experiment with reconstituted nucleosomes left some doubt on the interpretation due to its high exonuclease activity. Furthermore, MNase cleavage could not provide direct evidence of the structural state. This study used Förster resonance energy transfer (FRET) measurement to investigate the salt-induced conformational change of the nucleosomal DNA containing AA/TT repeat at the entry site. We observed that the AA/TT region wrapped around the histone core was as likely as other DNA sequences under the physiological salt concentration. However, it unwrapped at a lower salt concentration, indicating weaker electrostatic interactions with the histone core. The ethidium-induced nucleosome disruption assay showed that the intercalator had more access to the DNA with AA/TT at the entry site. Taken together, we suggest that AA/TT at the entry sites induces DNA unwraping from the histone core in the promoter side, promoting transcription activation in response to the approach of the transcription-related proteins.