The role of histone H4 biotinylation in the structure of nucleosomes.
Post-translational modifications of histones play important roles in regulating nucleosome structure and gene transcription. It has been shown that biotinylation of histone H4 at lysine-12 in histone H4 (K12Bio-H4) is associated with repression of a number of genes. We hypothesized that biotinylation modifies the physical structure of nucleosomes, and that biotin-induced conformational changes contribute to gene silencing associated with histone biotinylation.To test this hypothesis we used atomic force microscopy to directly analyze structures of nucleosomes formed with biotin-modified and non-modified H4. The analysis of the AFM images revealed a 13% increase in the length of DNA wrapped around the histone core in nucleosomes with biotinylated H4. This statistically significant (p<0.001) difference between native and biotinylated nucleosomes corresponds to adding approximately 20 bp to the classical 147 bp length of nucleosomal DNA.The increase in nucleosomal DNA length is predicted to stabilize the association of DNA with histones and therefore to prevent nucleosomes from unwrapping. This provides a mechanistic explanation for the gene silencing associated with K12Bio-H4. The proposed single-molecule AFM approach will be instrumental for studying the effects of various epigenetic modifications of nucleosomes, in addition to biotinylation.
PubMed ID: 21298003
PMC ID: PMC3029316
Article link: PLoS One.
Grant support: DK063945 NIDDK NIH HHS , DK077816 NIDDK NIH HHS , DK082476 NIDDK NIH HHS , P30CA036727 NCI NIH HHS
Genes referenced: hist1h4d
Article Images: [+] show captions
|Figure 1. Schematics for various stages of the nucleosome unwrapping.Nucleosome conformations are shown with different number of turns, rotation angle and length of wrapped DNA.|
|Figure 2. Representative AFM scans of nucleosome core particles.Nucleosomes were reconstituted using native histone H4 (a) or biotinylated histone K12Cbio-H4 (b). Images were acquired with NanoScope IIId AFM system operating in Tapping mode. Scan sizes are 0.5 µm.|
|Figure 3. Representative enlarged AFM scans of NCP.Nucleosomes were reconstituted using non-biotinylated native histone H4 (a) and biotinylated K12C histone H4 (b). The complexes are labeled with the number of DNA turns around histone octamers. Scans sizes are 200×200 nm.|
|Figure 4. Histograms for lengths of nucleosomal DNA (nsDNA) wrapped around histone cores.Nucleosomes were reconstituted using native H4 histone (a), K12C-H4 mutant (b) or K12Cbio-H4 histone (c). It can be seen that in nucleosomes made with K12Cbio-H4 wDNA is shifted towards higher value compared to samples reconstituted using non-biotinylated native H4 or K12C-H4 mutant. The mean values of nsDNA indicated with arrows were 49.8 nm ±1.5 nm, 48.8 nm ±1.4 nm and 56.6 nm ±1.1 nm for NCP containing native histone H4, K12C-H4 mutant, and K12Cbio-H4, respectively.|
|Figure 5. Model of the effect of biotinylation on conformation of nucleosome.Both front and top views are shown. The segment of the DNA arm that contributes to additional wrapping is shown in red.|