Date Published: January 5, 2017
Publisher: AIMS Press
Author(s): Daniel Sultanov, Nadezhda Gerasimova, Kseniya Kudryashova, Natalya Maluchenko, Elena Kotova, Marie-France Langelier, John Pascal, Mikhail Kirpichnikov, Alexey Feofanov, Vasily Studitsky.
DNA accessibility to various protein complexes is essential for various processes in the cell and is affected by nucleosome structure and dynamics. Protein factor PARP-1 (poly(ADP-ribose) polymerase 1) increases the accessibility of DNA in chromatin to repair proteins and transcriptional machinery, but the mechanism and extent of this chromatin reorganization are unknown. Here we report on the effects of PARP-1 on single nucleosomes revealed by spFRET (single-particle Förster Resonance Energy Transfer) microscopy. PARP-1 binding to a double-strand break in the vicinity of a nucleosome results in a significant increase of the distance between the adjacent gyres of nucleosomal DNA. This partial uncoiling of the entire nucleosomal DNA occurs without apparent loss of histones and is reversed after poly(ADP)-ribosylation of PARP-1. Thus PARP-1-nucleosome interactions result in reversible, partial uncoiling of the entire nucleosomal DNA.
Eukaryotic genome is composed of nucleosomes that consist of 145–148 bp DNA segments wrapped around the histone octamer in 1.65–1.7 superhelical coils. Nucleosomal organization limits DNA accessibility to various proteins, including protein complexes involved in DNA repair . Various protein complexes, including ATP-dependent chromatin remodelers and PARP-1 protein reorganize chromatin, making it more accessible to other DNA-interacting proteins.
Our spFRET experiments suggest that PARP-1 binds to a nucleosome and induces disturbance of different regions of nucleosomal DNA: near the entrance/exit of DNA into/from a nucleosome, and in the region positioned ∼35 bp from the boundaries of nucleosomal DNA (Figures 2 and 3). This uncoiling of nucleosomal DNA is accompanied by a reduced mobilities of the +13, +35 and +112 regions of nucleosomal DNA (Figures 2 and 3). PARP-1 automodification (self-PARylation) is accompanied by formation of an intermediate complex, and eventually leads to nearly complete recovery of the initial structure of nucleosome (Figure 4). Thus spFRET is a sensitive method for analysis of PARP-1-induced changes in chromatin structure that could also be used for analysis of PARP-1 inhibition by various compounds.
Nucleosome structure can be considerably and reversibly unfolded after PARP-1 binding. These changes include transient and partial uncoiling of nucleosomal DNA along its entire length. These PARP-1-dependent changes in nucleosome structure are nearly completely reversed after PARP-1 eviction due to its auto-poly(ADP)-ribosylation.