(2022). Aneuploidy tolerance caused by BRG1 loss allows chromosome gains and recovery of fitness. Nature communications,
Aneuploidy results in decreased cellular fitness in many species and model systems. However, aneuploidy is commonly found in cancer cells and often correlates with aggressive growth, suggesting that the impact of aneuploidy on cellular fitness is context dependent. The BRG1 (SMARCA4) subunit of the SWI/SNF chromatin remodelling complex is frequently lost in cancer. Here, we use a chromosomally stable cell line to test the effect of BRG1 loss on the evolution of aneuploidy. BRG1 deletion leads to an initial loss of fitness in this cell line that improves over time. Notably, we find increased tolerance to aneuploidy immediately upon loss of BRG1, and the fitness recovery over time correlates with chromosome gain. These data show that BRG1 loss creates an environment where karyotype changes can be explored without a fitness penalty. At least in some genetic backgrounds, therefore, BRG1 loss can affect the progression of tumourigenesis through tolerance of aneuploidy..
Del Nery, E.
(2021). PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer. Cancer research,
Inactivation of Polybromo 1 (PBRM1), a specific subunit of the PBAF chromatin remodeling complex, occurs frequently in cancer, including 40% of clear cell renal cell carcinomas (ccRCC). To identify novel therapeutic approaches to targeting PBRM1-defective cancers, we used a series of orthogonal functional genomic screens that identified PARP and ATR inhibitors as being synthetic lethal with PBRM1 deficiency. The PBRM1/PARP inhibitor synthetic lethality was recapitulated using several clinical PARP inhibitors in a series of in vitro model systems and in vivo in a xenograft model of ccRCC. In the absence of exogenous DNA damage, PBRM1-defective cells exhibited elevated levels of replication stress, micronuclei, and R-loops. PARP inhibitor exposure exacerbated these phenotypes. Quantitative mass spectrometry revealed that multiple R-loop processing factors were downregulated in PBRM1-defective tumor cells. Exogenous expression of the R-loop resolution enzyme RNase H1 reversed the sensitivity of PBRM1-deficient cells to PARP inhibitors, suggesting that excessive levels of R-loops could be a cause of this synthetic lethality. PARP and ATR inhibitors also induced cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) innate immune signaling in PBRM1-defective tumor cells. Overall, these findings provide the preclinical basis for using PARP inhibitors in PBRM1-defective cancers.
This study demonstrates that PARP and ATR inhibitors are synthetic lethal with the loss of PBRM1, a PBAF-specific subunit, thus providing the rationale for assessing these inhibitors in patients with PBRM1-defective cancer.
von Loga, K.
(2021). Differential and longitudinal immune gene patterns associated with reprogrammed microenvironment and viral mimicry in response to neoadjuvant radiotherapy in rectal cancer. Journal for immunotherapy of cancer,
Rectal cancers show a highly varied response to neoadjuvant radiotherapy/chemoradiation (RT/CRT) and the impact of the tumor immune microenvironment on this response is poorly understood. Current clinical tumor regression grading systems attempt to measure radiotherapy response but are subject to interobserver variation. An unbiased and unique histopathological quantification method (change in tumor cell density (ΔTCD)) may improve classification of RT/CRT response. Furthermore, immune gene expression profiling (GEP) may identify differences in expression levels of genes relevant to different radiotherapy responses: (1) at baseline between poor and good responders, and (2) longitudinally from preradiotherapy to postradiotherapy samples. Overall, this may inform novel therapeutic RT/CRT combination strategies in rectal cancer.Methods
We generated GEPs for 53 patients from biopsies taken prior to preoperative radiotherapy. TCD was used to assess rectal tumor response to neoadjuvant RT/CRT and ΔTCD was subjected to k-means clustering to classify patients into different response categories. Differential gene expression analysis was performed using statistical analysis of microarrays, pathway enrichment analysis and immune cell type analysis using single sample gene set enrichment analysis. Immunohistochemistry was performed to validate specific results. The results were validated using 220 pretreatment samples from publicly available datasets at metalevel of pathway and survival analyses.Results
ΔTCD scores ranged from 12.4% to −47.7% and stratified patients into three response categories. At baseline, 40 genes were significantly upregulated in poor (n=12) versus good responders (n=21), including myeloid and stromal cell genes. Of several pathways showing significant enrichment at baseline in poor responders, epithelial to mesenchymal transition, coagulation, complement activation and apical junction pathways were validated in external cohorts. Unlike poor responders, good responders showed longitudinal (preradiotherapy vs postradiotherapy samples) upregulation of 198 immune genes, reflecting an increased T-cell-inflamed GEP, type-I interferon and macrophage populations. Longitudinal pathway analysis suggested viral-like pathogen responses occurred in post-treatment resected samples compared with pretreatment biopsies in good responders.Conclusion
This study suggests potentially druggable immune targets in poor responders at baseline and indicates that tumors with a good RT/CRT response reprogrammed from immune “cold” towards an immunologically “hot” phenotype on treatment with radiotherapy..
(2021). Histone isoforms and the oncohistone code. Current opinion in genetics & development,
(2021). Improved RAD51 binders through motif shuffling based on the modularity of BRC repeats. Proceedings of the national academy of sciences,
Despite advances in directed evolution and computational design, engineering of functional proteins remains challenging. Here we demonstrate that rearrangement of modular repeats from BRCA2 can yield chimeras with improved interaction properties, while deconstructing the relative energetic contributions of different modules across a large interaction interface and resolving the effects of shape complementary or physicochemical properties by empirical observation. This de- and reconstruction of binding interfaces supports a mix-and-match model for proteins in which repeat units can be manipulated and used to construct functional proteins that interfere with dsDNA repair. Our approach may aid in creating biochemical and therapeutic tools from natural modules, with minimal screening effort..
(2020). The role of the SWI/SNF chromatin remodelling complex in the response to DNA double strand breaks. Dna repair (amst),
Mammalian cells possess multiple closely related SWI/SNF chromatin remodelling complexes. These complexes have been implicated in the cellular response to DNA double strand breaks (DSBs). Evidence suggests that SWI/SNF complexes contribute to successful repair via both the homologous recombination and non-homologous end joining pathways. In addition, repressing transcription near DSBs is dependent on SWI/SNF activity. Understanding these roles is important because SWI/SNF complexes are frequently dysregulated in cancer, and DNA DSB repair defects have the potential to be therapeutically exploited. In this graphical review, we summarise what is known about SWI/SNF contribution to DNA DSB responses in mammalian cells and provide an overview of the SWI/SNF-encoding gene alteration spectrum in human cancers..
van Attikum, H.
(2020). CHD7 and 53BP1 regulate distinct pathways for the re-ligation of DNA double-strand breaks. Nature communications,
Chromatin structure is dynamically reorganized at multiple levels in response to DNA double-strand breaks (DSBs). Yet, how the different steps of chromatin reorganization are coordinated in space and time to differentially regulate DNA repair pathways is insufficiently understood. Here, we identify the Chromodomain Helicase DNA Binding Protein 7 (CHD7), which is frequently mutated in CHARGE syndrome, as an integral component of the non-homologous end-joining (NHEJ) DSB repair pathway. Upon recruitment via PARP1-triggered chromatin remodeling, CHD7 stimulates further chromatin relaxation around DNA break sites and brings in HDAC1/2 for localized chromatin de-acetylation. This counteracts the CHD7-induced chromatin expansion, thereby ensuring temporally and spatially controlled ‘chromatin breathing’ upon DNA damage, which we demonstrate fosters efficient and accurate DSB repair by controlling Ku and LIG4/XRCC4 activities. Loss of CHD7-HDAC1/2-dependent cNHEJ reinforces 53BP1 assembly at the damaged chromatin and shifts DSB repair to mutagenic NHEJ, revealing a backup function of 53BP1 when cNHEJ fails..
(2019). Repression of Transcription at DNA Breaks Requires Cohesin throughout Interphase and Prevents Genome Instability. Molecular cell,
(2017). Chromatin modifiers and remodellers in DNA repair and signalling. Philosophical transactions of the royal society b: biological sciences,
(2016). Novel synthetic lethality screening method identifies TIP60-dependent radiation sensitivity in the absence of BAF180. Dna repair,
(2016). Discovery and Optimization of a Selective Ligand for the Switch/Sucrose Nonfermenting-Related Bromodomains of Polybromo Protein-1 by the Use of Virtual Screening and Hydration Analysis. Journal of medicinal chemistry,
(2015). The PBAF chromatin remodeling complex represses transcription and promotes rapid repair at DNA double-strand breaks. Molecular & cellular oncology,
(2015). Removal of H2A Z by
80 promotes homologous recombination. Embo reports,
(2015). The SWI/SNF chromatin remodelling complex: Its role in maintaining genome stability and preventing tumourigenesis. Dna repair,
(2015). The BAH domain of BAF180 is required for PCNA ubiquitination. Mutation research/fundamental and molecular mechanisms of mutagenesis,
(2015). Fork rotation and DNA precatenation are restricted during DNA replication to prevent chromosomal instability. Proceedings of the national academy of sciences,
Genome inheritance requires the complete resolution of all intertwines within parental DNA. This is facilitated by fork rotation and precatenation of the newly replicated DNA. However, the general importance and frequency of fork rotation in vivo are poorly understood. We find that the evolutionarily conserved Timeless and Tipin proteins actively inhibit fork rotation in budding yeast. In their presence, fork rotation appears restricted to hard-to-replicate fragile sites. In their absence, excessive fork rotation leads to damage accumulating in the replicated sister chromatids, especially at known yeast fragile sites. Therefore, fork rotation appears to be restricted to contexts where it is absolutely required for unwinding, and this restriction is required to prevent precatenation inducing excessive chromosomal fragility..
(2014). BAF180 Promotes Cohesion and Prevents Genome Instability and Aneuploidy. Cell reports,
(2014). Structural Basis for Phosphorylation-Dependent Recruitment of Tel2 to Hsp90 by Pih1. Structure,
(2014). Structure of Yin Yang 1 Oligomers That Cooperate with RuvBL1-RuvBL2 ATPases. Journal of biological chemistry,
(2014). Requirement for PBAF in Transcriptional Repression and Repair at DNA Breaks in Actively Transcribed Regions of Chromatin. Molecular cell,
(2014). Roles of chromatin remodellers in DNA double strand break repair. Experimental cell research,
(2013). The BAH domain of Rsc2 is a histone H3 binding domain. Nucleic acids research,
(2012). A role for chromatin remodellers in replication of damaged DNA. Nucleic acids research,
(2012). Cancer and the bromodomains of BAF180. Biochemical society transactions,
Chromatin remodelling complexes alter the structure of chromatin and have central roles in all DNA-templated activities, including regulation of gene expression and DNA repair. Mutations in subunits of the PBAF (polybromo/Brg1-associated factor) or SWI/SNF-B remodelling complex, including BAF180, are frequently associated with cancer. There are six potential acetyl-lysine-binding BDs (bromodomains) in BAF180, which may function to target the PBAF complex to promoters or sites of DNA repair. In the present review, we discuss what is currently known about the BDs of BAF180 and their potential significance in cancer..
(2012). The INO80 chromatin remodeling complex prevents polyploidy and maintains normal chromatin structure at centromeres. Genes & development,
The INO80 chromatin remodeling complex functions in transcriptional regulation, DNA repair, and replication. Here we uncover a novel role for INO80 in regulating chromosome segregation. First, we show that the conserved Ies6 subunit is critical for INO80 function in vivo. Strikingly, we found that loss of either Ies6 or the Ino80 catalytic subunit results in rapid increase in ploidy. One route to polyploidy is through chromosome missegregation due to aberrant centromere structure, and we found that loss of either Ies6 or Ino80 leads to defective chromosome segregation. Importantly, we show that chromatin structure flanking centromeres is altered in cells lacking these subunits and that these alterations occur not in the Cse4-containing centromeric nucleosome, but in pericentric chromatin. We provide evidence that these effects are mediated through misincorporation of H2A.Z, and these findings indicate that H2A.Z-containing pericentric chromatin, as in higher eukaryotes with regional centromeres, is important for centromere function in budding yeast. These data reveal an important additional mechanism by which INO80 maintains genome stability..
(2012). The RSC and INO80 Chromatin-Remodeling Complexes in DNA Double-Strand Break Repair. ,
(2009). Methylation of H3 K4 and K79 is not strictly dependent on H2B K123 ubiquitylation. Journal of cell biology,
Covalent modifications of histone proteins have profound consequences on chromatin structure and function. Specific modification patterns constitute a code read by effector proteins. Studies from yeast found that H3 trimethylation at K4 and K79 is dependent on ubiquitylation of H2B K123, which is termed a “trans-tail pathway.” In this study, we show that a strain unable to be ubiquitylated on H2B (K123R) is still proficient for H3 trimethylation at both K4 and K79, indicating that H3 methylation status is not solely dependent on H2B ubiquitylation. However, additional mutations in H2B result in loss of H3 methylation when combined with htb1-K123R. Consistent with this, we find that the original strain used to identify the trans-tail pathway has a genomic mutation that, when combined with H2B K123R, results in defective H3 methylation. Finally, we show that strains lacking the ubiquitin ligase Bre1 are defective for H3 methylation, suggesting that there is an additional Bre1 substrate that in combination with H2B K123 facilitates H3 methylation..
(2009). A metabolomic and multivariate statistical process to assess the effects of genotoxins in Saccharomycescerevisiae. Molecular biosystems,
(2008). Histone H3 K56 acetylation, chromatin assembly, and the DNA damage checkpoint. Dna repair,
(2007). The contribution of the budding yeast histone H2A C-terminal tail to DNA-damage responses. Biochemical society transactions,
The cellular response to DNA damage involves extensive interaction with and manipulation of chromatin. This includes the detection and repair of the DNA lesion, but there are also transcriptional responses to DNA damage, involving the up- or down-regulation of numerous genes. Therefore changes to chromatin structure, including covalent modification of histone proteins, are known to occur during DNA-damage responses. One of the most well characterized DNA-damage-responsive chromatin modification events is the phosphorylation of the SQ motif found in the C-terminal tail of histone H2A or the H2AX variant in higher eukaryotes. In the budding yeast, a number of additional residues in this region of histone H2A that contribute to the cellular response to DNA damage have been identified, providing an insight into the nature and complexity of the DNA-damage histone code..
(2007). Dual Chromatin Remodeling Roles for RSC during DNA Double Strand Break Induction and Repair at the Yeast MAT Locus. Journal of biological chemistry,
(2007). Chromatin structure and DNA double-strand break responses in cancer progression and therapy. Oncogene,
(2007). The Bre5/Ubp3 ubiquitin protease complex from budding yeast contributes to the cellular response to DNA damage. Dna repair,
(2007). Chromatin dynamics and the preservation of genetic information. Nature,
(2005). Tails of histones in DNA double-strand break repair. Mutagenesis,
(2005). Saccharomyces cerevisiae Histone H2A Ser122 Facilitates DNA Repair. Genetics,
DNA repair takes place in the context of chromatin. Recently, it has become apparent that proteins that make up and modulate chromatin structure are involved in the detection and repair of DNA lesions. We previously demonstrated that Ser129 in the carboxyl-terminal tail of yeast histone H2A is important for double-strand-break responses. By undertaking a systematic site-directed mutagenesis approach, we identified another histone H2A serine residue (Ser122) that is important for survival in the presence of DNA-damaging agents. We show that mutation of this residue does not affect DNA damage-dependent Rad53 phosphorylation or G2/M checkpoint responses. Interestingly, we find that yeast lacking H2A S122 are defective in their ability to sporulate. Finally, we demonstrate that H2A S122 provides a function distinct from that of H2A S129. These data demonstrate a role for H2A S122 in facilitating survival in the presence of DNA damage and suggest a potential role in mediating homologous recombination. The distinct roles of H2A S122 and S129 in mediating these responses suggest that chromatin components can provide specialized functions for distinct DNA repair and survival mechanisms and point toward the possibility of a complex DNA damage responsive histone code..
(2005). Histones as tumour suppressor genes. Cellular and molecular life sciences,
(2005). Spotting new DNA damage-responsive chromatin-binding proteins. Biochemical journal,
In response to DNA damage, cells initiate multiple repair mechanisms that all contribute to the survival of both the cell and the organism. These responses are numerous and variable, and can include cell cycle arrest, transcriptional activation of DNA repair genes and relocalization of repair proteins to sites of DNA damage. If all else fails, in multicellular organisms the initiation of apoptosis is also a potential cellular response to DNA damage. Despite a wealth of information about these events, it is clear that we do not yet have a comprehensive picture of the cellular responses to DNA damage. In this issue of the Biochemical Journal, a proteomics approach was used by Lee et al. to identify proteins that bind to chromatin in a DNA damage-inducible manner. The proteins identified, nucleophosmin, hnRNP C1 (heterogeneous nuclear ribonucleoprotein C1) and hnRNP C2, were proteins that would not necessarily have been predicted to behave this way. These studies have the potential to be extended and contribute to our knowledge of the cellular response to DNA damage..
(2005). Dynamics of Chromatin during the Repair of DNA Double-Strand Breaks. Cell cycle,
(2004). A means to a DNA end: the many roles of Ku. Nature reviews molecular cell biology,
(2004). Binding of Chromatin-Modifying Activities to Phosphorylated Histone H2A at DNA Damage Sites. Molecular cell,
(2003). Suppression of Homologous Recombination by the Saccharomyces cerevisiae Linker Histone. Molecular cell,
(2002). Interdependent Interactions between TFIIB, TATA Binding Protein, and DNA. Molecular and cellular biology,
Temperature-sensitive mutants of TFIIB that are defective for essential interactions were isolated. One mutation (G204D) results in disruption of a protein-protein contact between TFIIB and TATA binding protein (TBP), while the other (K272I) disrupts an interaction between TFIIB and DNA. The TBP gene was mutagenized, and alleles that suppress the slow-growth phenotypes of the TFIIB mutants were isolated. TFIIB with the G204D mutation [TFIIB(G204D)] was suppressed by hydrophobic substitutions at lysine 239 of TBP. These changes led to increased affinity between TBP and TFIIB. TFIIB(K272I) was weakly suppressed by TBP mutants in which K239 was changed to hydrophobic residues. However, this mutant TFIIB was strongly suppressed by conservative substitutions in the DNA binding surface of TBP. Biochemical characterization showed that these TBP mutants had increased affinity for a TATA element. The TBPs with increased affinity could not suppress TFIIB(G204D), leading us to propose a two-step model for the interaction between TFIIB and the TBP-DNA complex..
(2000). A role for Saccharomyces cerevisiae histone H2A in DNA repair. Nature,
(1999). Involvement of DNA End-Binding Protein Ku in Ty Element Retrotransposition. Molecular and cellular biology,
Ty elements are retrotransposons whose life cycles are strikingly similar to those of retroviruses. They transpose via an RNA intermediate that is converted to linear double-stranded cDNA and then inserted into the host genome. Although Ty integration is mediated by the element-encoded integrase, it has been proposed that host factors are involved in this process. Here, we show that the DNA end-binding protein Ku, which functions in DNA double-strand break repair, potentiates retrotransposition. Specifically, by using a galactose-inducible Ty1 system, we found that in vivo, Ty1 retrotransposition rates were substantially reduced in the absence of Ku. In contrast, this phenotype was not observed with yeast strains containing mutations in other genes that are involved in DNA repair. We present evidence that Ku associates with Ty1 viruslike particles both in vitro and in vivo. These results provide an additional role for Ku and suggest that it might function in the life cycles of retroelements in other systems.
Epigenetic changes in histone acetylation underpin resistance to the topoisomerase I inhibitor irinotecan. Nucleic acids research,
Predicting synthetic lethal interactions using conserved patterns in protein interaction networks. Plos computational biology,
Histone H2A phosphorylation in DNA double-strand break repair. Febs journal,
What functions do linker histones provide?. Molecular microbiology,
The Two Different Isoforms of the RSC Chromatin Remodeling Complex Play Distinct Roles in DNA Damage Responses. Plos one,
TP53 MODULATES RADIOTHERAPY FRACTION SIZE SENSITIVITY IN NORMAL AND MALIGNANT CELLS. Scientific reports,
Defining Signatures of Arm-Wise Copy Number Change and Their Associated Drivers in Kidney Cancers. International journal of molecular sciences,
Using pan-cancer data from The Cancer Genome Atlas (TCGA), we investigated how patterns in copy number alterations in cancer cells vary both by tissue type and as a function of genetic alteration. We find that patterns in both chromosomal ploidy and individual arm copy number are dependent on tumour type. We highlight for example, the significant losses in chromosome arm 3p and the gain of ploidy in 5q in kidney clear cell renal cell carcinoma tissue samples. We find that specific gene mutations are associated with genome-wide copy number changes. Using signatures derived from non-negative factorisation, we also find gene mutations that are associated with particular patterns of ploidy change. Finally, utilising a set of machine learning classifiers, we successfully predicted the presence of mutated genes in a sample using arm-wise copy number patterns as features. This demonstrates that mutations in specific genes are correlated and may lead to specific patterns of ploidy loss and gain across chromosome arms. Using these same classifiers, we highlight which arms are most predictive of commonly mutated genes in kidney renal clear cell carcinoma (KIRC)..