Chromatin remodeling is the dynamic modification of chromatin architecture to allow access of condensed genomic DNA to the regulatory transcription machinery proteins, and thereby control gene expression. Such remodeling is mainly carried out by 1) covalent histone modifications by specific enzymes, e.g., histone acetyltransferases (HATs), histone deacetylases (HDACs), methyltransferases, and kinases, and 2) ATP-dependent chromatin remodeling complexes which either move, eject or restructure nucleosomes. Besides actively regulating gene expression, dynamic remodeling of chromatin imparts an epigenetic regulatory role in several key biological processes, egg cells DNA replication and repair; apoptosis; chromosome segregation as well as development and pluripotency. Aberrations in chromatin remodeling proteins are found to be associated with human diseases, including cancer. Targeting chromatin remodeling pathways is currently evolving as a major therapeutic strategy in the treatment of several cancers.
Overview
The transcriptional regulation of the genome is controlled primarily at the preinitiation stage by binding of the core transcriptional machinery proteins (namely, RNA polymerase, transcription factors, activators and repressors) to the core promoter sequence on the coding region of the DNA. However, DNA is tightly packaged in the nucleus with the help of packaging proteins, chiefly histone proteins to form repeating units of nucleosomes which further bundle together to form condensed chromatin structure. Such condensed structure occludes many DNA regulatory regions, not allowing them to interact with transcriptional machinery proteins and regulate gene expression. To overcome this issue and allow dynamic access to condensed DNA, a process known as chromatin remodeling alters nucleosome architecture to expose or hide regions of DNA for transcriptional regulation.
By definition, chromatin remodeling is the enzyme-assisted process to facilitate access of nucleosomal DNA by remodeling the structure, composition and positioning of nucleosomes.
Classification Access to nucleosomal DNA is governed by two major classes of protein complexes: Covalent histone-modifying complexes. ATP-dependent chromatin remodeling complexes.
Covalent histone-modifying complexes Specific protein complexes, known as histone-modifying complexes catalyze addition or removal of various chemical elements on histones. These enzymatic modifications include acetylation, methylation, phosphorylation, ubiquitination, and citrullination and primarily occur at N-terminal histone tails. Such modifications affect the binding affinity between histones and DNA, and thus loosening or tightening the condensed DNA wrapped around histones, e.g., methylation of specific lysine residues in H3 and H4 causes further condensation of DNA around histones, and thereby prevents binding of transcription factors (TF) to the DNA that lead to gene repression. On the contrary, histone acetylation relaxes chromatin condensation and exposes DNA for TF binding, leading to increased gene expression.
Known modifications Well characterized modifications to histones include: Methylation Both lysine and arginine residues are known to be methylated. Methylated lysines are the best understood marks of the histone code, as specific methylated lysine match well with gene expression states. Methylation of lysines H3K4 and H3K36 is correlated with transcriptional activation while demethylation of H3K4 is correlated with silencing of the genomic region. Methylation of lysines H3K9 and H3K27 is correlated with transcriptional repression. Particularly, H3K9me3 is highly correlated with constitutive heterochromatin. Acetylation - by HAT (histone acetyl transferase); deacetylation - by HDAC (histone deacetylase) Acetylation tends to define the 'openness' of chromatin as acetylated histones cannot pack as well together as deacetylated histones. Phosphorylation Ubiquitination Citrullination However, there are many more histone modifications, and sensitive mass spectrometry approaches have recently greatly expanded the catalog.
Histone code hypothesis The histone code is a hypothesis that the transcription of genetic information encoded in DNA is in part regulated by chemical modifications to histone proteins, primarily on their unstructured ends. Together with similar modifications such as DNA methylation it is part of the epigenetic code.
Cumulative evidence suggests that such code is written by specific enzymes which can (for example) methylate or acetylate histones ('writers'), removed by other enzymes having demethylase or deacetylase activity ('erasers'), and finally readily identified by proteins ('readers') that are recruited to such histone modifications and bind via specific domains, e.g., bromodomain, chromodomain. These triple action of 'writing', 'reading' and 'erasing' establish the favorable local environment for transcriptional regulation, DNA-damage repair, etc.
The critical concept of the histone code hypothesis is that the histone modifications serve to recruit other proteins by specific recognition of the modified histone via protein domains specialized for such purposes, rather than through simply stabilizing or destabilizing the interaction between histone and the underlying DNA. These recruited proteins then act to alter chromatin structure actively or to promote transcription.
A very basic summary of the histone code for gene expression status is given below (histone nomenclature is described here):
Type ofmodification Histone H3K4 H3K9 H3K14 H3K27 H3K79 H4K20 H2BK5 mono-methylation activation activation activation activation activation activation di-methylation repression repression activation tri-methylation activation repression repression activation,repression repression acetylation activation activation
ATP-dependent chromatin remodeling
ATP-dependent chromatin-remodeling complexes regulate gene expression by either moving, ejecting or restructuring nucleosomes. These protein complexes have a common ATPase domain and energy from the hydrolysis of ATP allows these remodeling complexes to reposition nucleosomes (often referred to as "nucleosome sliding") along the DNA, eject or assemble histones on/off of DNA or facilitate exchange of histone variants, and thus creating nucleosome-free regions of DNA for gene activation. Also, several remodelers have DNA-translocation activity to carry out specific remodeling tasks.
All ATP-dependent chromatin-remodeling complexes possess a sub unit of ATPase that belongs to the SNF2 superfamily of proteins. In association to the sub unit's identity, two main groups have been classified for these proteins. These are known as the SWI2/SNF2 group and the imitation SWI (ISWI) group. The third class of ATP-dependent complexes that has been recently described contains a Snf2-like ATPase and also demonstrates deacetylase activity.
Known chromatin remodeling complexes
There are at least four families of chromatin remodelers in eukaryotes: SWI/SNF, ISWI, NuRD/Mi-2/CHD, and INO80 with first two remodelers being very well studied so far, especially in the yeast model. Although all of remodelers share common ATPase domain, their functions are specific based on several biological processes (DNA repair, apoptosis, etc.). This is due to the fact that each remodeler complex has unique protein domains (Helicase, bromodomain, etc.) in their catalytic ATPase region and also has different recruited subunits.
Specific functions Several in-vitro experiments suggest that ISWI remodelers organize nucleosome into proper bundle form and create equal spacing between nucleosomes, whereas SWI/SNF remodelers disorder nucleosomes. The ISWI-family remodelers have been shown to play central roles in chromatin assembly after DNA replication and maintenance of higher-order chromatin structures. INO80 and SWI/SNF-family remodelers participate in DNA double-strand break (DSB) repair and nucleotide-excision repair (NER) and thereby plays crucial role in TP53 mediated DNA-damage response. NuRD/Mi-2/CHD remodeling complexes primarily mediate transcriptional repression in the nucleus and are required for the maintenance of pluripotency of embryonic stem cells.
Significance
In normal biological processes Chromatin remodeling plays a central role in the regulation of gene expression by providing the transcription machinery with dynamic access to an otherwise tightly packaged genome. Further, nucleosome movement by chromatin remodelers is essential to several important biological processes, including chromosome assembly and segregation, DNA replication and repair, embryonic development and pluripotency, and cell-cycle progression. Deregulation of chromatin remodeling causes loss of transcriptional regulation at these critical check-points required for proper cellular functions, and thus causes various disease syndromes, including cancer.
Response to DNA damage Chromatin relaxation is one of the earliest cellular responses to DNA damage. Several experiments have been performed on the recruitment kinetics of proteins involved in the response to DNA damage. The relaxation appears to be initiated by PARP1, whose accumulation at DNA damage is half complete by 1.6 seconds after DNA damage occurs. This is quickly followed by accumulation of chromatin remodeler Alc1, which has an ADP-ribose–binding domain, allowing it to be quickly attracted to the product of PARP1. The maximum recruitment of Alc1 occurs within 10 seconds of DNA damage. About half of the maximum chromatin relaxation, presumably due to action of Alc1, occurs by 10 seconds. PARP1 action at the site of a double-strand break allows recruitment of the two DNA repair enzymes MRE11 and NBS1. Half maximum recruitment of these two DNA repair enzymes takes 13 seconds for MRE11 and 28 seconds for NBS1.
Another process of chromatin relaxation, after formation of a DNA double-strand break, employs γH2AX, the phosphorylated form of the H2AX protein. The histone variant H2AX constitutes about 10% of the H2A histones in human chromatin. γH2AX (phosphorylated on serine 139 of H2AX) was detected at 20 seconds after irradiation of cells (with DNA double-strand break formation), and half maximum accumulation of γH2AX occurred in one minute. The extent of chromatin with phosphorylated γH2AX is about two million base pairs at the site of a DNA double-strand break.
γH2AX does not, by itself, cause chromatin decondensation, but within seconds of irradiation the protein "Mediator of the DNA damage checkpoint 1" (MDC1) specifically attaches to γH2AX. This is accompanied by simultaneous accumulation of RNF8 protein and the DNA repair protein NBS1 which bind to MDC1 as MDC1 attaches to γH2AX. RNF8 mediates extensive chromatin decondensation, through its subsequent interaction with CHD4 protein, a component of the nucleosome remodeling and deacetylase complex NuRD. CHD4 accumulation at the site of the double-strand break is rapid, with half-maximum accumulation occurring by 40 seconds after irradiation.
The fast initial chromatin relaxation upon DNA damage (with rapid initiation of DNA repair) is followed by a slow recondensation, with chromatin recovering a compaction state close to its pre-damage level in ~ 20 min.
Cancer Chromatin remodeling provides fine-tuning at crucial cell growth and division steps, like cell-cycle progression, DNA repair and chromosome segregation, and therefore exerts tumor-suppressor function. Mutations in such chromatin remodelers and deregulated covalent histone modifications potentially favor self-sufficiency in cell growth and escape from growth-regulatory cell signals - two important hallmarks of cancer.