title: Nucleosomal stability and dynamics vary significantly when viewed by internal versus terminal labels.
abstract: Nucleosomes are a major impediment to regulatory factor activities and therefore to the operation of genomic processes in eukaryotes. One suggested mechanism for overcoming in vivo nucleosomal repression is factor-mediated removal of H2A/H2B from nucleosomes. Using nucleosomes labeled internally with FRET fluorophores, we previously observed significant, DNA sequence-dependent variation in stability and dynamics under conditions (subnanomolar concentrations) reported to produce H2A/H2B release from nucleosomes. Here, the same analytical approaches are repeated using 5S and MMTV-B nucleosomes containing FRET labels that monitor the terminal regions. The results show that stability and dynamics vary significantly within the nucleosome; terminally labeled constructs report significantly reduced stability and enhanced DNA dynamics compared to internally labeled constructs. The data also strongly support previous suggestions (1) that subnanomolar concentrations cause H2A/H2B release from nucleosomes, including the 5S, and (2) that stabilities in the internal regions of 5S and two promoter-derived nucleosomes (MMTV-B, GAL10) differ. Sequence-dependent nucleosome stability/dynamics differences could produce inherent variations in the accessibility of histone-associated DNA in vivo. Such intrinsic variation could also provide a mechanism for producing enhanced effects on specific nucleosomes by processes affecting large chromatin regions, thus facilitating the localized targeting of alterations to nucleosomes on crucial regulatory sequences. The results demonstrate clearly the importance of studying physiologically relevant nucleosomes.
authors.affiliation: Biodesign Institute, Arizona State University, Tempe, Arizona 85287, USA. laimonas.kelbauskas@asu.edu
authors.forename: Laimonas
authors.initials: L
authors.lastname: Kelbauskas
authors.forename: Jenny
authors.initials: J
authors.lastname: Sun
authors.forename: Neal
authors.initials: N
authors.lastname: Woodbury
authors.forename: D
authors.initials: D
authors.lastname: Lohr
mesh_terms: D002474:Cell-Free System / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry* / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry*; D006367:HeLa Cells; D006657:Histones / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008324:Mammary Tumor Virus, Mouse / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009707:Nucleosomes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D004279:DNA, Viral; D005456:Fluorescent Dyes; C518355:GAL10 protein, S cerevisiae; D006657:Histones; D009707:Nucleosomes; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators