Chromatin Assembly Factor Research Articles

HP1 cooperates with CAF-1 to compact heterochromatic transgene repeats in mammalian cells

The nuclear organization of tightly condensed heterochromatin plays important roles in regulating gene transcription and genome integrity. Heterochromatic domains are usually present at chromosomal regions containing a large array of repeated DNA sequences. We previously showed that integration of a 1,000-copy tandem array of an inducible reporter gene into the genome of mammalian cells induces the formation of a highly compact heterochromatic domain enriched in heterochromatin protein 1 (HP1). It remains to be determined how these DNA repeats are packaged into a heterochromatic form and are silenced. Here, we show that HP1-mediated transgene condensation and silencing require the interaction with PxVxL motif-containing proteins. The chromatin assembly factor 1 (CAF-1) complex concentrates at the transgenic locus through the interaction of its PxVxL motif-containing p150 subunit with HP1. Knockdown of p150 relieves HP1-mediated transgene compaction and repression. When targeted to the transgenic locus, p150 mutants defective in binding HP1 cause transgene decondensation and activation. Taken together, these results suggest that HP1 cooperates with CAF-1 to compact transgene repeats. This study provides important insight into how heterochromatin is maintained at chromosomal regions with abundant DNA repeats.

Mechanistic insights into histone deposition and nucleosome assembly by the chromatin assembly factor-1.

Eukaryotic chromatin is a highly dynamic structure with essential roles in virtually all DNA-dependent cellular processes. Nucleosomes are a barrier to DNA access, and during DNA replication, they are disassembled ahead of the replication machinery (the replisome) and reassembled following its passage. The Histone chaperone Chromatin Assembly Factor-1 (CAF-1) interacts with the replisome and deposits H3–H4 directly onto newly synthesized DNA. Therefore, CAF-1 is important for the establishment and propagation of chromatin structure. The molecular mechanism by which CAF-1 mediates H3–H4 deposition has remained unclear. However, recent studies have revealed new insights into the architecture and stoichiometry of the trimeric CAF-1 complex and how it interacts with and deposits H3–H4 onto substrate DNA. The CAF-1 trimer binds to a single H3–H4 dimer, which induces a conformational rearrangement in CAF-1 promoting its interaction with substrate DNA. Two CAF-1•H3–H4 complexes co-associate on nucleosome-free DNA depositing (H3–H4)2 tetramers in the first step of nucleosome assembly. Here, we review the progress made in our understanding of CAF-1 structure, mechanism of action, and how CAF-1 contributes to chromatin dynamics during DNA replication.

The replicative histone chaperone CAF1 is essential for the maintenance of identity and genome integrity in adult stem cells.

Chromatin packaging and modifications are important to define the identity of stem cells. How chromatin properties are retained over multiple cycles of stem cell replication, while generating differentiating progeny at the same time, remains a challenging question. The chromatin assembly factor CAF1 is a conserved histone chaperone, which assembles histones H3 and H4 onto newly synthesized DNA during replication and repair. Here, we have investigated the role of CAF1 in the maintenance of germline stem cells (GSCs) in Drosophila ovaries. We depleted P180, the large subunit of CAF1, in germ cells and found that it was required in GSCs to maintain their identity. In the absence of P180, GSCs still harbor stem cell properties but concomitantly express markers of differentiation. In addition, P180-depleted germ cells exhibit elevated levels of DNA damage and de-repression of the transposable I element. These DNA damages activate p53- and Chk2-dependent checkpoints pathways, leading to cell death and female sterility. Altogether, our work demonstrates that chromatin dynamics mediated by CAF1 play an important role in both the regulation of stem cell identity and genome integrity.

The histone chaperone ASF1 regulates the activation of ATM and DNA-PKcs in response to DNA double-strand breaks

ABSTRACTThe Ataxia-telangiectasia mutated (ATM) kinase and the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) are activated by DNA double-strand breaks (DSBs). These DSBs occur in the context of chromatin but how chromatin influences the activation of these kinases is not known. Here we show that loss of the replication-dependent chromatin assembly factors ASF1A/B or CAF-1 compromises ATM activation, while augmenting DNA-PKcs activation, in response to DNA DSBs. Cells deficient in ASF1A/B or CAF-1 exhibit reduced histone H4 lysine 16 acetylation (H4K16ac), a histone mark known to promote ATM activation. ASF1A interacts with the histone acetyl transferase, hMOF that mediates H4K16ac. ASF1A depletion leads to increased recruitment of DNA-PKcs to DSBs. We propose normal chromatin assembly and H4K16ac during DNA replication is required to regulate ATM and DNA-PKcs activity in response to the subsequent induction of DNA DSBs.

Replication‐coupled histone H3.1 deposition determines nucleosome composition and heterochromatin dynamics during Arabidopsis seedling development

Developmental phase transitions are often characterized by changes in the chromatin landscape and heterochromatin reorganization. In Arabidopsis, clustering of repetitive heterochromatic loci into so-called chromocenters is an important determinant of chromosome organization in nuclear space. Here, we investigated the molecular mechanisms involved in chromocenter formation during the switch from a heterotrophic to a photosynthetically competent state during early seedling development. We characterized the spatial organization and chromatin features at centromeric and pericentromeric repeats and identified mutant contexts with impaired chromocenter formation. We find that clustering of repetitive DNA loci into chromocenters takes place in a precise temporal window and results in reinforced transcriptional repression. Although repetitive sequences are enriched in H3K9me2 and linker histone H1 before repeat clustering, chromocenter formation involves increasing enrichment in H3.1 as well as H2A.W histone variants, hallmarks of heterochromatin. These processes are severely affected in mutants impaired in replication-coupled histone assembly mediated by CHROMATIN ASSEMBLY FACTOR 1 (CAF-1). We further reveal that histone deposition by CAF-1 is required for efficient H3K9me2 enrichment at repetitive sequences during chromocenter formation. Taken together, we show that chromocenter assembly during post-germination development requires dynamic changes in nucleosome composition and histone post-translational modifications orchestrated by the replication-coupled H3.1 deposition machinery.

Nucleosomes around a mismatched base pair are excluded via an Msh2-dependent reaction with the aid of SNF2 family ATPase Smarcad1.

Post-replicative correction of replication errors by the mismatch repair (MMR) system is critical for suppression of mutations. Although the MMR system may need to handle nucleosomes at the site of chromatin replication, how MMR occurs in the chromatin environment remains unclear. Here, we show that nucleosomes are excluded from a >1-kb region surrounding a mismatched base pair in Xenopus egg extracts. The exclusion was dependent on the Msh2-Msh6 mismatch recognition complex but not the Mlh1-containing MutL homologs and counteracts both the HIRA- and CAF-1 (chromatin assembly factor 1)-mediated chromatin assembly pathways. We further found that the Smarcad1 chromatin remodeling ATPase is recruited to mismatch-carrying DNA in an Msh2-dependent but Mlh1-independent manner to assist nucleosome exclusion and that Smarcad1 facilitates the repair of mismatches when nucleosomes are preassembled on DNA. In budding yeast, deletion of FUN30, the homolog of Smarcad1, showed a synergistic increase of spontaneous mutations in combination with MSH6 or MSH3 deletion but no significant increase with MSH2 deletion. Genetic analyses also suggested that the function of Fun30 in MMR is to counteract CAF-1. Our study uncovers that the eukaryotic MMR system has an ability to exclude local nucleosomes and identifies Smarcad1/Fun30 as an accessory factor for the MMR reaction.

Histone chaperones play crucial roles in maintenance of stem cell niche during plant root development.

Stem cells in both plant and animal kingdoms reside in a specialized cellular context called the stem cell niche (SCN). SCN integrity is crucial for organism development. Here we show that the H3/H4 histone chaperone CHROMATIN ASSEMBLY FACTOR-1 (CAF-1) and the H2A/H2B histone chaperone NAP1-RELATED PROTEIN1/2 (NRP1/2) play synergistic roles in Arabidopsis root SCN maintenance. Compared with either the m56-1 double mutant deprived of NRP1 and NRP2 or the fas2-4 mutant deprived of CAF-1, the combined m56-1fas2-4 triple mutant displayed a much more severe short-root phenotype. The m56-1fas2-4 mutant root lost the normal organizing center Quiescent Center (QC), and some initial stem cells differentiated precociously. Microarray analysis unraveled the deregulation of 2735 genes within the Arabidopsis genome (representing >8% of all genes) in the m56-1fas2-4 mutant roots. Expression of some SCN key regulatory genes (e.g. WOX5, PLT1, SHR) was not limiting, rather the plant hormone auxin gradient maximum at QC was impaired. The mutant roots showed programmed cell death and high levels of the DNA damage marked histone H2A.X phosphorylation (γ-H2A.X). Knockout of either ATAXIA-TELANGIECTASIA MUTATED (ATM) or ATR, encoding a DNA damage response kinase, rescued in part the cell death and the short-root phenotype of the m56-1fas2-4 mutant. Collectively, our study indicated that NRP1/2 and CAF-1 act cooperatively in regulating proper genome transcription, in sustaining chromatin replication and in maintaining genome integrity, which are crucial for proper SCN function during continuous post-embryonic root development.

Chromatin assembly factor-1 (CAF-1) chaperone regulates Cse4 deposition into chromatin in budding yeast.

Correct localization of the centromeric histone variant CenH3/CENP-A/Cse4 is an important part of faithful chromosome segregation. Mislocalization of CenH3 could affect chromosome segregation, DNA replication and transcription. CENP-A is often overexpressed and mislocalized in cancer genomes, but the underlying mechanisms are not understood. One major regulator of Cse4 deposition is Psh1, an E3 ubiquitin ligase that controls levels of Cse4 to prevent deposition into non-centromeric regions. We present evidence that Chromatin assembly factor-1 (CAF-1), an evolutionarily conserved histone H3/H4 chaperone with subunits shown previously to interact with CenH3 in flies and human cells, regulates Cse4 deposition in budding yeast. yCAF-1 interacts with Cse4 and can assemble Cse4 nucleosomes in vitro. Loss of yCAF-1 dramatically reduces the amount of Cse4 deposited into chromatin genome-wide when Cse4 is overexpressed. The incorporation of Cse4 genome-wide may have multifactorial effects on growth and gene expression. Loss of yCAF-1 can rescue growth defects and some changes in gene expression associated with Cse4 deposition that occur in the absence of Psh1-mediated proteolysis. Incorporation of Cse4 into promoter nucleosomes at transcriptionally active genes depends on yCAF-1. Overall our findings suggest CAF-1 can act as a CenH3 chaperone, regulating levels and incorporation of CenH3 in chromatin.

Transgenerational phenotype aggravation in CAF-1 mutants reveals parent-of-origin specific epigenetic inheritance.

Chromatin is assembled by histone chaperones such as chromatin assembly factor CAF-1. We had noticed that vigor of Arabidopsis thaliana CAF-1 mutants decreased over several generations. Because changes in mutant phenotype severity over generations are unusual, we asked how repeated selfing of Arabidopsis CAF-1 mutants affects phenotype severity. CAF-1 mutant plants of various generations were grown, and developmental phenotypes, transcriptomes and DNA cytosine-methylation profiles were compared quantitatively. Shoot- and root-related growth phenotypes were progressively more affected in successive generations of CAF-1 mutants. Early and late generations of the fasciata (fas)2-4 CAF-1 mutant displayed only limited changes in gene expression, of which increasing upregulation of plant defense-related genes reflects the transgenerational phenotype aggravation. Likewise, global DNA methylation in the sequence context CHG but not CG or CHH (where H = A, T or C) changed over generations in fas2-4. Crossing early and late generation fas2-4 plants established that the maternal contribution to the phenotype severity exceeds the paternal contribution. Together, epigenetic rather than genetic mechanisms underlie the progressive developmental phenotype aggravation in the Arabidopsis CAF-1 mutants and preferred maternal transmission reveals a more efficient reprogramming of epigenetic information in the male than the female germline.

The development of functional mapping by three sex-related loci on the third whorl of different sex types of Carica papaya L.

Carica papaya L. is an important economic crop worldwide and is used as a model plant for sex-determination research. To study the different flower sex types, we screened sex-related genes using alternative splicing sequences (AS-seqs) from a transcriptome database of the three flower sex types, i.e., males, females, and hermaphrodites, established at 28 days before flowering using 15 bacterial artificial chromosomes (BACs) of C. papaya L. After screening, the cDNA regions of the three sex-related loci, including short vegetative phase-like (CpSVPL), the chromatin assembly factor 1 subunit A-like (CpCAF1AL), and the somatic embryogenesis receptor kinase (CpSERK), which contained eight sex-related single-nucleotide polymorphisms (SNPs) from the different sex types of C. papaya L., were genotyped using high-resolution melting (HRM). The three loci were examined regarding the profiles of the third whorl, as described below. CpSVPL, which had one SNP associated with the three sex genotypes, was highly expressed in the male and female sterile flowers (abnormal hermaphrodite flowers) that lacked the fourth whorl structure. CpCAF1AL, which had three SNPs associated with the male genotype, was highly expressed in male and normal hermaphrodite flowers, and had no AS-seqs, whereas it exhibited low expression and an AS-seqs in intron 11 in abnormal hermaphrodite flowers. Conversely, carpellate flowers (abnormal hermaphrodite flowers) showed low expression of CpSVPL and AS-seqs in introns 5, 6, and 7 of CpSERK, which contained four SNPs associated with the female genotype. Specifically, the CpSERK and CpCAF1AL loci exhibited no AS-seq expression in the third whorl of the male and normal hermaphrodite flowers, respectively, and variance in the AS-seq expression of all other types of flowers. Functional mapping of the third whorl of normal hermaphrodites indicated no AS-seq expression in CpSERK, low CpSVPL expression, and, for CpCAF1AL, high expression and no AS-seq expression on XYh-type chromosomes.

NuRD and CAF-1-mediated silencing of the D4Z4 array is modulated by DUX4-induced MBD3L proteins.

The DUX4 transcription factor is encoded by a retrogene embedded in each unit of the D4Z4 macrosatellite repeat. DUX4 is normally expressed in the cleavage-stage embryo, whereas chromatin repression prevents DUX4 expression in most somatic tissues. Failure of this repression causes facioscapulohumeral muscular dystrophy (FSHD) due to mis-expression of DUX4 in skeletal muscle. In this study, we used CRISPR/Cas9 engineered chromatin immunoprecipitation (enChIP) locus-specific proteomics to characterize D4Z4-associated proteins. These and other approaches identified the Nucleosome Remodeling Deacetylase (NuRD) and Chromatin Assembly Factor 1 (CAF-1) complexes as necessary for DUX4 repression in human skeletal muscle cells and induced pluripotent stem (iPS) cells. Furthermore, DUX4-induced expression of MBD3L proteins partly relieved this repression in FSHD muscle cells. Together, these findings identify NuRD and CAF-1 as mediators of DUX4 chromatin repression and suggest a mechanism for the amplification of DUX4 expression in FSHD muscle cells.

Chromatin assembly factor-1 (CAF-1) chaperone regulates Cse4 deposition into chromatin in budding yeast.

Correct localization of the centromeric histone variant CenH3/CENP-A/Cse4 is an important part of faithful chromosome segregation. Mislocalization of CenH3 could affect chromosome segregation, DNA replication and transcription. CENP-A is often overexpressed and mislocalized in cancer genomes, but the underlying mechanisms are not understood. One major regulator of Cse4 deposition is Psh1, an E3 ubiquitin ligase that controls levels of Cse4 to prevent deposition into non-centromeric regions. We present evidence that Chromatin assembly factor-1 (CAF-1), an evolutionarily conserved histone H3/H4 chaperone with subunits shown previously to interact with CenH3 in flies and human cells, regulates Cse4 deposition in budding yeast. yCAF-1 interacts with Cse4 and can assemble Cse4 nucleosomes in vitro. Loss of yCAF-1 dramatically reduces the amount of Cse4 deposited into chromatin genome-wide when Cse4 is overexpressed. The incorporation of Cse4 genome-wide may have multifactorial effects on growth and gene expression. Loss of yCAF-1 can rescue growth defects and some changes in gene expression associated with Cse4 deposition that occur in the absence of Psh1-mediated proteolysis. Incorporation of Cse4 into promoter nucleosomes at transcriptionally active genes depends on yCAF-1. Overall our findings suggest CAF-1 can act as a CenH3 chaperone, regulating levels and incorporation of CenH3 in chromatin.

Identification of PCNA Binding Sites on Chromatin Assembly Factor 1

Identification of PCNA Binding Sites on Chromatin Assembly Factor 1

Structural Basis of Heterochromatin Formation by Human HP1

Heterochromatin plays important roles in transcriptional silencing and genome maintenance by the formation of condensed chromatin structures, which determine the epigenetic status of eukaryotic cells.The trimethylation of histone H3 lysine 9 (H3K9me3), a target of heterochromatin protein 1 (HP1), is a hallmark of heterochromatin formation. However, the mechanism by which HP1 folds chromatin-containing H3K9me3 into a higher-order structure has not been elucidated. Here we report the three-dimensional structure of the H3K9me3-containing dinucleosomes complexed with human HP1α, HP1β, and HP1γ, determined by cryogenic electron microscopy with a Volta phase plate. In the structures, two H3K9me3 nucleosomes are bridged by a symmetric HP1 dimer. Surprisingly, the linker DNA between the nucleosomes does not directly interact with HP1, thus allowing nucleosome remodeling by the ATP-utilizing chromatin assembly and remodeling factor (ACF). The structure depicts the fundamental architecture of heterochromatin.

A genetic link between epigenetic repressor AS1–AS2 and DNA replication factors in establishment of adaxial–abaxial leaf polarity of <i>Arabidopsis</i>

Balanced development of adaxial and abaxial domains in leaf primordia is critical for the formation of flat symmetric leaf lamina. Arabidopsis ASYMMETRIC LEAVES1 (AS1) and AS2 proteins form a complex (AS1-AS2), which acts as key regulators for the adaxial development by the direct repression of expression of the abaxial gene ETTIN/AUXIN RESPONSE FACTOR3 (ETT/ARF3). Many modifier mutations have been identified, which enhance the defect of as1 and as2 mutations to generate abaxialized filamentous leaves without adaxial traits, suggesting that the development of the adaxial domain is achieved by cooperative repression by AS1-AS2 and the wild-type proteins corresponding to the modifiers. Mutations of several genes for DNA replication-related chromatin remodeling factors such as Chromatin Assembly Factor-1 (CAF-1) have been also identified as modifiers. It is still unknown, however, whether mutations in genes involved in DNA replication themselves might act as modifiers. Here we report that as1 and as2 mutants grown in the presence of hydroxyurea, a known inhibitor of DNA replication, form abaxialized filamentous leaves in a concentration-dependent manner. We further show that a mutation of the INCURVATA2 (ICU2) gene, which encodes the putative catalytic subunit of DNA polymerase α, and a mutation of the Replication Factor C Subunit3 (RFC3) gene, which encodes a protein used in replication as a clamp loader, act as modifiers. In addition, as2-1 icu2-1 double mutants showed increased mRNA levels of the genes for leaf abaxialization. These results suggest a tight link between DNA replication and the function of AS1-AS2 in the development of flat leaves.

CHRAC/ACF contribute to the repressive ground state of chromatin.

The chromatin remodeling complexes chromatin accessibility complex and ATP-utilizing chromatin assembly and remodeling factor (ACF) combine the ATPase ISWI with the signature subunit ACF1. These enzymes catalyze well-studied nucleosome sliding reactions in vitro, but how their actions affect physiological gene expression remains unclear. Here, we explored the influence of Drosophila melanogaster chromatin accessibility complex/ACF on transcription by using complementary gain- and loss-of-function approaches. Targeting ACF1 to multiple reporter genes inserted at many different genomic locations revealed a context-dependent inactivation of poorly transcribed reporters in repressive chromatin. Accordingly, single-embryo transcriptome analysis of an Acf knock-out allele showed that only lowly expressed genes are derepressed in the absence of ACF1. Finally, the nucleosome arrays in Acf-deficient chromatin show loss of physiological regularity, particularly in transcriptionally inactive domains. Taken together, our results highlight that ACF1-containing remodeling factors contribute to the establishment of an inactive ground state of the genome through chromatin organization.

Role of Chromatin assembly factor 1 in DNA replication of Plasmodium falciparum

Nucleosome assembly in P. falciparum could be the key process in maintaining its genomic integrity as DNA replicates more than once per cell cycle during several stages of its life cycle. Here, we report the functional characterization of P. falciparum chromatin assembly factor 1 (CAF1), which interacts with several proteins namely PfCAF2, Histones, PfHP1 and others. Consistent with the above findings, we demonstrate the presence of PfCAF1 at the telomeric repeat regions, central and subtelomeric var genes of multiple var gene family along with PfHP1. Further, we report the upregulation of PfCAF1 after treatment with genotoxic agents like MMS and HU. Together, these findings establish role of PfCAF1 in heterochromatin maintenance and as histone chaperone in nucleosome assembly and DNA damage repair.

CHAF1A, the largest subunit of the chromatin assembly factor?1 complex, regulates the growth of H1299 human non‑small cell lung cancer cells by inducing G0/G1 cell cycle arrest

Chromatin assembly factor 1 subunit A (CHAF1A) is the largest subunit of the chromatin assembly factor 1 (CAF-1) complex that is implicated in the assembly of nucleosomes on newly synthesized DNA. The aim of the present study was to determine its expression and biological function in non-small cell lung cancer (NSCLC). The current study examined the levels of CHAF1A expression in 22 samples of NSCLC and corresponding normal lung tissues. Subsequently, endogenous CHAF1A expression in H1299 NSCLC cells was knocked down via lentiviral delivery of CHAF1A-targeting short hairpin RNA (shRNA), and cell proliferation, colony formation and cell cycle distribution were measured. The results demonstrated that levels of CHAF1A mRNA level were ~3-fold greater in NSCLC samples compared with adjacent normal tissues (P<0.05). shRNA-mediated silencing of CHAF1A significantly inhibited the proliferation and colony formation of H1299 cells, compared wirh the delivery of control shRNA (P<0.05). Furthermore, CHAF1A shRNA-transduced cells exhibited a significant increase in the percentage of S-phase cells and a significant decrease in the percentage of cells at the G0/G1 and G2/M phases, compared with control cells (P<0.05). Additionally, CHAF1A knockdown significantly decreased the expression of cyclin D1, cyclin-dependent kinase 2 and S-phase kinase-associated protein 2, and increased the expression of p21 and p27. This indicates that CHAF1A is upregulated in NSCLC and that its silencing suppresses the proliferation and colony formation of NSCLC cells, potentially by inducing G0/G1 cell cycle arrest. CHAF1A may therefore represent a potential therapeutic target to treat NSCLC.

Arabidopsis Chromatin Assembly Factor 1 is required for occupancy and position of a subset of nucleosomes.

Chromatin Assembly Factor 1 (CAF-1) is a major nucleosome assembly complex which functions particularly during DNA replication and repair. Here we studied how the nucleosome landscape changes in a CAF-1 mutant in the model plant Arabidopsis thaliana. Globally, most nucleosomes were not affected by loss of CAF-1, indicating the presence of efficient alternative nucleosome assemblers. Nucleosomes that we found depleted in the CAF-1 mutant were enriched in non-transcribed regions, consistent with the notion that CAF-1-independent nucleosome assembly can compensate for loss of CAF-1 mainly in transcribed regions. Depleted nucleosomes were particularly enriched in proximal promoters, suggesting that CAF-1-independent nucleosome assembly mechanisms are often not efficient upstream of transcription start sites. Genes related to plant defense were particularly prone to lose nucleosomes in their promoters upon CAF-1 depletion. Reduced nucleosome occupancy at promoters of many defense-related genes is associated with a primed gene expression state that may considerably increase plant fitness by facilitating plant defense. Together, our results establish that the nucleosome landscape in Arabidopsis is surprisingly robust even in the absence of the dedicated nucleosome assembly machinery CAF-1 and that CAF-1-independent nucleosome assembly mechanisms are less efficient in particular genome regions.

Nucleosome remodelling, DNA repair and transcriptional regulation build negative feedback loops in cancer and cellular ageing.

Nucleosome remodelling (NR) regulates transcription in an ATP-dependent manner, and influences gene expression required for development and cellular functions, including those involved in anti-cancer and anti-ageing processes. ATP-utilizing chromatin assembly and remodelling factor (ACF) and Brahma-associated factor (BAF) complexes, belonging to the ISWI and SWI/SNF families, respectively, are involved in various types of DNA repair. Suppression of several BAF factors makes U2OS cells significantly sensitive to X-rays, UV and especially to cisplatin, and these BAF factors contribute to the accumulation of repair proteins at various types of DNA damage and to DNA repair. Recent cancer genome sequencing and expression analysis has shown that BAF factors are frequently mutated or, more frequently, silenced in various types of cancer cells. Thus, those cancer cells are potentially X-ray- and especially cisplatin-sensitive, suggesting a way of optimizing current cancer therapy. Recent single-stem cell analysis suggests that mutations and epigenetic changes influence stem cell functionality leading to cellular ageing. Genetic and epigenetic changes in the BAF factors diminish DNA repair as well as transcriptional regulation activities, and DNA repair defects in turn negatively influence NR and transcriptional regulation. Thus, they build negative feedback loops, which accelerate both cellular senescence and transformation as common and rare cellular events, respectively, causing cellular ageing.This article is part of the themed issue 'Chromatin modifiers and remodellers in DNA repair and signalling'.