Nucleosome traversal by RNA polymerase II (pol II) and recovery of chromatin structure after transcription are essential for proper gene expression. In this paper we show that nucleosomes assembled with Sin mutant histones present a much weaker barrier to traversal by pol II and are less likely to survive transcription. Increases in traversal from incorporation of Sin mutant histones and histones lacking H2A/H2B amino-terminal tails were in most cases additive, indicating that traversal can be facilitated by distinct mechanisms. We had identified a key intermediate in traversal, the zero (slashed circle)-loop, which mediates nucleosome survival during transcription. Sin mutations probably destabilize these intermediates and thus increase the likelihood of nucleosome disassociation.
"Genetic studies found that either a glutamic acid or alanine substitution for this residue is lethal in haploid yeast, while low-level expression of H3(T118E) and H3(T118A) resulted in defects in transcriptional silencing (27). In addition, H3(T118I) is a SWI/SNF INdependent (SIN) mutation, where this substitution relieves the requirement of the SWItch/Sucrose NonFermentable (SWI/SNF) chromatin remodeling complex for mating type switching in budding yeast (28), and either mutation H3(T118I) (29) or H3(T118H) (30) lowers the transcriptional barrier for Pol II. Our recent studies of single nucleosomes containing H3(T118ph) found that this phosphorylation dramatically enhanced nucleosome mobility, reduced DNA–histone binding and facilitated nucleosome disassembly by SWI/SNF (20). "
[Show abstract][Hide abstract] ABSTRACT: Nucleosomes contain ∼146 bp of DNA wrapped around a histone protein octamer that controls DNA accessibility to transcription and repair complexes. Posttranslational modification (PTM) of histone proteins regulates nucleosome function. To date, only modest changes in nucleosome structure have been directly attributed to histone PTMs. Histone residue H3(T118) is located near the nucleosome dyad and can be phosphorylated. This PTM destabilizes nucleosomes and is implicated in the regulation of transcription and repair. Here, we report gel electrophoretic mobility, sucrose gradient sedimentation, thermal disassembly, micrococcal nuclease digestion and atomic force microscopy measurements of two DNA-histone complexes that are structurally distinct from nucleosomes. We find that H3(T118ph) facilitates the formation of a nucleosome duplex with two DNA molecules wrapped around two histone octamers, and an altosome complex that contains one DNA molecule wrapped around two histone octamers. The nucleosome duplex complex forms within short ∼150 bp DNA molecules, whereas altosomes require at least ∼250 bp of DNA and form repeatedly along 3000 bp DNA molecules. These results are the first report of a histone PTM significantly altering the nucleosome structure.
Nucleic Acids Research 02/2014; 42(8). DOI:10.1093/nar/gku150 · 9.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Creating, maintaining, or using a digital library requires the manipulation of digital documents. Information workspaces provide a visual representation allowing users to collect, organize, annotate, and author information. The visual knowledge builder (VKB) helps users access, collect, annotate, and combine materials from digital libraries and other sources into a personal information workspace. VKB has been enhanced to include direct search interfaces for NSDL and Google. Users create a visualization of search results while selecting and organizing materials for their current activity. Additionally, metadata applicators have been added to VKB. This interface allows the rapid addition of metadata to documents and aids the user in the extraction of existing metadata for application to other documents. A study was performed to compare the selection and organization of documents in VKB to the commonly used tools of a Web browser and a word processor. This study shows the value of visual workspaces for such effort but points to the need for subdocument level objects, ephemeral visualizations, and support for moving from visual representations to metadata.
ACM/IEEE Joint Conference on Digital Libraries, JCDL 2004, Tucson, AZ, USA, June 7-11, 2004, Proceedings; 01/2004
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