Yes / Topoisomerase1 (TOP1)-mediated chromosomal breaks are endogenous sources of DNA damage that affect neuronal genome stability. Whether TOP1 DNA breaks are sources of genomic instability in Huntington's disease (HD) is unknown. Here, we report defective 53BP1 recruitment in multiple HD cell models, including striatal neurons derived from HD patients. Defective 53BP1 recruitment is due to reduced H2A ubiquitination caused by the limited RNF168 activity. The reduced availability of RNF168 is caused by an increased interaction with p62, a protein involved in selective autophagy. Depletion of p62 or disruption of the interaction between RNAF168 and p62 was sufficient to restore 53BP1 enrichment and subsequent DNA repair in HD models, providing new opportunities for therapeutic interventions. These findings are reminiscent to what was described for p62 accumulation caused by C9orf72 expansion in ALS/FTD and suggest a common mechanism by which protein aggregation perturb DNA repair signaling. / This work is funded by a Welcome Trust Investigator Award (103844), a Lister Institute of Preventative Medicine Fellowship (137661) and a UKIERI grant (DST/INT/UK/P-147/2016) to S.F.E.- K. JG is additionally funded by a Clinical PhD Fellowship from the Pathological Society of Great Britain and Ireland and the Jean Shanks Foundation (JSPS-CPHD-2018-01).

Eukaryotic organisms contain their entire genome in the nucleus of their cells. In order to fit within the nucleus, genomic DNA wraps into nucleosomes, the basic, repeating unit of chromatin. Nucleosomes wrap around each other to form higher order chromatin structures. Here we study many factors that affect, or are effected by, chromatin structure including: (1) how low-dose inorganic arsenic (iAs) changes chromatin structures and their relation to global transcription and splicing patterns, and (2) how chromatin architectural proteins (CAPs) bind to and change nucleosome dynamics and DNA target site accessibility. Despite iAs’s non-mutagenic nature, chronic exposure to low doses of iAs is associated with a higher risk of skin, lung, and bladder cancers. We sought to identify the genome-wide changes to chromatin structure and splicing profiles behind the cell’s adaptive response to iAs and its removal. Furthermore, we extended our investigation into cells that had the iAs insult removed. Our results show that the iAs-induced epithelial to mesenchymal transition and changes to the transcriptome are coupled with changes to the higher order chromatin structure and CAP binding patterns. We hypothesize that CAPs, which bind the entry/exit and linker DNA of nucleosomes, regulate DNA target site accessibility by altering of the rate of spontaneous dissociation of DNA from nucleosome. Therefore, we investigated the effects of the repressive CAP histone H1, the activating CAP high mobility group D1 (HMGD1), and the neural CAP methyl CpG binding protein 2 (MeCP2) on the dynamics of short chromatin arrays and mononucleosomes and their effect on nucleosomal DNA accessibility. Using biochemical and biophysical analyses we show that all CAP-chromatin structures tested were susceptible to chromatin remodeling by ISWI and created more stable higher order structures than if CAPs were absent. Additionally, histone H1 and MeCP2 hinder model transcription factor Gal4 from binding its cognate DNA site within nucleosomal DNA. Overall, we show that chromatin structure is dynamic and changes in response to environmental signals and that CAPs change nucleosome dynamics that help to regulate chromatin structures and impact transcriptional profiles.

Der pRB/E2F-Signalweg ist ein zentraler Regulator der Proliferationskontrolle in Säugerzellen, der in fast allen auftretenden Tumoren dereguliert ist. Durch unterschiedliche Mutationen in Komponenten dieses Signalwegs kommt es letzten Endes zu einer erhöhten Aktivität der E2F-Transkriptionsfaktoren und somit zu einer verstärkten Transkription von E2F-Zielgenen in diesen Tumoren. Um die molekularen Mechanismen der Rolle von E2F3 in der Zellzykluskontrolle und der Tumorigenese besser zu verstehen, wurden in dieser Arbeit per GST-Pulldown mit anschließender Massenspektrometrie neue potenzielle Interaktions-partner von E2F3 identifiziert. Ein identifizierter Interaktionspartner war die SNF2-ähnliche Helikase HELLS. HELLS interagiert in vitro und in vivo spezifisch mit der Marked Box-Domäne von E2F3, aber nicht mit anderen untersuchten E2F-Transkriptionsfaktoren, wie durch GST-Interaktionsstudien und Ko-Immunpräzipi-tationsexperimente demonstriert werden konnte. Durch Chromatin-Immunpräzipitation konnte zusätzlich gezeigt werden, dass E2F3 für die Rekrutierung von HELLS an E2F-regulierte Promotoren wie z. B. CDC6 oder p107 verantwortlich ist. Die shRNA-vermittelte Depletion von HELLS führte zu einer stark verminderten Induktion von allen untersuchten E2F-Zielgenen nach Serumstimulation und einem verspäteten Eintritt in die S-Phase der HELLS-depletierten Zellen, was zeigt, dass HELLS essenziell für die Induktion von E2F-Zielgenen ist. Bei der immunhistochemischen Untersuchung der E2F3- und HELLS-Expression in humanen Prostatakarzinomen zeigte sich, dass sowohl E2F3 als auch HELLS in späten aggressiven Stadien dieser Tumore sehr stark exprimiert sind, jedoch nur sehr schwach in den weniger aggressiven Tumoren. Diese Versuche zeigen, dass es sich bei HELLS um einen neuen Bestandteil des pRB/E2F-Signalwegs handelt, der eventuell in der Entstehung gewisser Tumorarten eine Rolle spielt und somit ein neues potenzielles Ziel für neuartige Krebstherapien darstellt. / The pRB/E2F pathway is a key regulator of proliferation in mammalian cells and is commonly mutated in human tumors. These mutations in the components of the pRB/E2F pathway lead to deregulated activity of the E2F transcription factors resulting in increased expression of E2F target genes. To further understand the molecular mechanisms of E2F3 in cell cycle control and its role in tumorigenesis new interaction partners for E2F3 were identified in the course of this thesis with the help of a GST-Pulldown approach coupled to mass spectrometric analysis. One of the identified interaction partners was the SNF2-like helicase HELLS. With the help of GST-interaction studies and Co-Immunoprecipitation assays it could be demonstrated that HELLS interacts specifically with E2F3 via its Marked Box domain but does not bind to the other investigated E2F transcription factors. HELLS could be detected at E2F target genes like p107 and CDC6 in vivo with the help of Chromatin-Immunoprecipitation assays. Furthermore, the forced recruitment of E2F3 to E2F target genes led to an enhanced binding of HELLS to these promotors suggesting that HELLS is recruited to E2F target genes via protein-protein interaction with E2F3. The shRNA-mediated depletion of HELLS led to a strongly reduced induction of E2F target genes and a delay in S-phase entry, showing that HELLS is essential for the induction of E2F target genes. During the immunohistochemical analysis of human prostate cancer specimens it became evident that both E2F3 and HELLS are strongly expressed in the more aggressive late stages but only weakly expressed in the early stages of this tumor type. These findings demonstrate that HELLS is a new component of the E2F/pRB pathway which might play a role in the development of certain tumors and might represent a new target for novel cancer therapies.

Mouse embryonic stem cells (mESCs) are an excellent model to study epigenetics and chromatin structure, owing to their self-renewal capabilities and tolerance of dynamic changes to DNA and histone modifications. Culturing conditions impact on the ability of mESCs to effectively recapitulate in vivo developmental states, and this is exemplified by refined culture conditions (termed 2i) that promote a pluripotent ground state. 2i-cultured mESC populations are homogeneous, naïve, and distinct from conventional (serum/LIF-cultured) cells, which exist as a metastable population. Remarkably, 2i-cultured mESCs also display global DNA hypomethylation, with methylation patterns more comparable to the cells of the E3.5 pre-implantation blastocyst. This is distinct from conventional serum-cultured cells, which display DNA methylation profiles that resemble later-stage E6.5 post-implantation epiblasts. The ability to transition between 2i- and serum-culture states is an attractive model for studying the dynamic role of DNA methylation in a variety of processes. DNA hypomethylation has been linked with depletion of the Polycomb-mediated repressive histone mark H3K27me3 from its normal target loci. Polycomb repressive complexes (PRC1 and PRC2) are important developmental regulators that maintain the repression of lineage-specific genes through generating compact higher-order chromatin structures. Polycomb target sites are primarily unmethylated CpG islands (CGIs). However, under conditions of DNA hypomethylation, new (previously methylated) binding sites are unveiled, and Polycomb is redistributed from its normal CGI target regions to intragenic regions. Thus, shifting mESCs to ground state conditions results in both DNA methylation and Polycomb patterns that are quite distinct from their serum-cultured counterparts. In my PhD, I sought to investigate the effect of DNA hypomethylation and Polycomb redistribution on higher-order chromatin structure in the ground state. I used a targeted, single-locus approach (FISH) as well as a genome-wide approach (Hi-C) to analyse differences in chromatin structure between conventionally cultured and ground state mESCs. My work suggests that chromatin structure is globally altered in hypomethylated 2icultured mESCs, with a similar state present in E3.5 mouse blastocysts. Using mESC lines in which DNA methylation levels can be directly manipulated, I was able to dissect the molecular mechanism driving higher-order structure changes in 2i medium, and showed the importance of DNA methylation in directing Polycomb-mediated chromatin compaction. My results may be important in considering the impact of DNA-methylation mediated reprogramming in multiple developmental, disease and regenerative medicine contexts.

CCCTC-binding factor (CTCF) binds DNA, thereby helping to partition the mammalian genome into discrete structural and regulatory domains. In doing so, it insulates chromatin and fine-tunes gene activation, repression, and silencing. Complete removal of CTCF from mammalian cells causes catastrophic genomic dysregulation, most likely due to widespread collapse of 3D chromatin looping within the nucleus. In contrast, Ctcf hemizygous mice with lifelong reduction in CTCF expression are viable but have an increased incidence of spontaneous multi-lineage malignancies. In addition, CTCF is mutated in many human cancers and is thus implicated as a tumour suppressor gene. This study aimed to interrogate the genome-wide consequences of a reduced genomic concentration of Ctcf and its implications for carcinogenesis. In a genetically engineered mouse model, Ctcf hemizygous cells showed modest but robust changes in almost a thousand sites of genomic CTCF occupancy; these were enriched for lower affinity binding events with weaker evolutionary conservation across the mouse lineage. Furthermore, several hundred genes concentrated in cancer-related pathways were dysregulated due to changes in transcriptional regulation. Global chromatin structure was preserved but some loop interactions were destabilised, often around differentially expressed genes and their enhancers. Importantly, these transcriptional alterations were also seen in human cancers. These findings were then examined in a hepatocyte-specific mouse model of Ctcf hemizygosity with diethylnitrosamine-induced liver tumours. Ctcf hemizygous mice had a subtle liver-specific phenotype, although the overall tumour burden in Ctcf hemizygous and wild-type mice was the same. Using whole genome sequencing, the highly reproducible mutational signature caused by DEN exposure was characterised, revealing that Braf(V637E), orthologous to BRAF(V600E) in humans, was the predominant oncogenic driver in these liver tumours. Taken together, while Ctcf loss is partially physiologically compensated, chronic CTCF depletion dysregulates gene expression by subtly altering transcriptional regulation. This study also represents the first comprehensive genome-wide and histopathological characterisation of this commonly used liver cancer model.

In eukaryotic cells, the genome is packed into chromatin, a hierarchically organized complex composed of DNA and histone and nonhistone proteins. In this thesis we have addressed the role of chromatin in cellular response to DNA damage (DDR) using various methodologies encompassing functional genomics and proteomics. First, we analyzed histone post-translational modifications (PTM) in the context of specific kind of DNA lesions (ICL-Interstrand Crosslinks) in Fanconi anemia using quantitative proteomics methodology, SILAC (Stable Isotope Labeling of Amino acids during Cell Culture). Using mass spectrometry (MS), we have successfully identified and quantified a number of histone PTM marks in histone H3 and H4, mainly acetylations and methylations,which have shown dependence upon functional FA-pathway. As a next step, we applied a functional genomics approach to study DDR in FA cells. In this analysis we first monitored the expression profile of histone modifying enzymes related to histone acetylations and methylations. Our results suggest some correlations between histone PTMs and gene expression of histone modifying enzymes, although conclusive evidence warrants further investigations. Next, we analyzed the total transcriptome after DNA damage induction in FA mutant and wild type cells. We also included in this analysis IR irradiation, in an attempt to dissociate more generic DDR from more specific changes that are associated with the role of FA pathway to the DNA ICLs. By performing a factorial interaction analysis, we were able to isolate the part of transcriptional response to DNA damage that was requiring functional FA pathway, as well as the genes that were sensitized to DNA damage by the inactivation of FA pathway. In the final part of the thesis, we attempted to solve one of the limitations that we encountered in the histone PTM analysis. The current approaches used to study histone PTMs from particular loci involves classical chromatin immunoprecipitation, which due to involvement of formaldehyde crosslinking render the protein part mostly unavailable for MS-based proteomics. We have proposed a novel methodology, which is based upon the biotin tagging of histones proximal to a protein of interest and subsequent purification of nucleosomes carrying the tagged histone. This methodology does not involve any crosslinking, enabling us to purify histones from specific loci, and subject them to large scale MS-based histone PTM analysis. A time dimension can also be added to our approach, as we can follow the modification status of particular fraction of histones once they get biotinylated. Another advantage is the use of alternate variant histones, which allows us to study the PTM profile of different functional states of chromatin. This methodology certainly has an edge on current techniques to study histone PTMs pattern associated with a particular protein of interest or with particular chromatin state.

No description available.

Transcription factors (TFs) are ubiquitous gene expression regulators and play essential roles in almost all biological processes. This Ph.D. project is primarily focused on the functional characterisation of MYB112 - a member of the R2R3-MYB TF family from the model plant Arabidopsis thaliana. This gene was selected due to its increased expression during senescence based on previous qRT-PCR expression profiling experiments of 1880 TFs in Arabidopsis leaves at three developmental stages (15 mm leaf, 30 mm leaf and 20% yellowing leaf). MYB112 promoter GUS fusion lines were generated to further investigate the expression pattern of MYB112. Employing transgenic approaches in combination with metabolomics and transcriptomics we demonstrate that MYB112 exerts a major role in regulation of plant flavonoid metabolism. We report enhanced and impaired anthocyanin accumulation in MYB112 overexpressors and MYB112-deficient mutants, respectively. Expression profiling reveals that MYB112 acts as a positive regulator of the transcription factor PAP1 leading to increased anthocyanin biosynthesis, and as a negative regulator of MYB12 and MYB111, which both control flavonol biosynthesis. We also identify MYB112 early responsive genes using a combination of several approaches. These include gene expression profiling (Affymetrix ATH1 micro-arrays and qRT-PCR) and transactivation assays in leaf mesophyll cell protoplasts. We show that MYB112 binds to an 8-bp DNA fragment containing the core sequence (A/T/G)(A/C)CC(A/T)(A/G/T)(A/C)(T/C). By electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation coupled to qPCR (ChIP-qPCR) we demonstrate that MYB112 binds in vitro and in vivo to MYB7 and MYB32 promoters revealing them as direct downstream target genes. MYB TFs were previously reported to play an important role in controlling flavonoid biosynthesis in plants. Many factors acting upstream of the anthocyanin biosynthesis pathway show enhanced expression levels during nitrogen limitation, or elevated sucrose content. In addition to the mentioned conditions, other environmental parameters including salinity or high light stress may trigger anthocyanin accumulation. In contrast to several other MYB TFs affecting anthocyanin biosynthesis pathway genes, MYB112 expression is not controlled by nitrogen limitation, or carbon excess, but rather is stimulated by salinity and high light stress. Thus, MYB112 constitutes a previously uncharacterised regulatory factor that modifies anthocyanin accumulation under conditions of abiotic stress. / Transkriptionsfaktoren (TFs) sind ubiquitäre Regulatoren der Genexpression und spielen eine essentielle Rolle in nahezu allen biologischen Prozessen. Diese Doktorarbeit hat vor allem die funktionelle Charakterisierung von MYB112 zum Thema, einem Mitglied der R2R3-MYB-TF-Familie aus der Modellpflanze Arabidopsis thaliana. Ausgesucht wurde das Gen aufgrund seiner erhöhten Expression in seneszenten Blättern, basierend auf vorangegangenen qRT-PCR Expressions-Profiling Experimenten für 1880 TFs in Arabidopsis Blättern aus drei Entwicklungsstadien (15 mm Blatt, 30 mm Blatt und 20 % vergilbtes Blatt). MYB112-Promotor-GUS-Fusionslinien wurden generiert um das Expressionsmuster von MYB112 detailliert zu untersuchen. Unter Zuhilfenahme transgener Ansätze in Kombination mit Metabolomics und Transcriptomics können wir zeigen, dass MYB112 eine wichtige Rolle in der Regulation des pflanzlichen Flavonoid-Metabolismus spielt. In MYB112 Überexpressoren und MYB112-defizienten Mutanten kommt es zu erhöhter bzw. verminderter Anthocyanin-Akkumulation. Expressions-Profiling zeigt, dass MYB112 einerseits als ein positiver Regulator des Transkriptionsfaktors PAP1 fungiert, was zu einer erhöhten Anthocyanin-Biosynthese führt, andererseits als negativer Regulator von MYB12 und MYB111 auftritt, welche beide die Flavonol-Biosynthese kontrollieren. Wir haben früh auf MYB112 reagierende Gene durch eine Kombination verschiedener Ansätze identifiziert. Diese umfassen Genexpressions-Profiling (Affymetrix ATH1 Microarrays und qRT-PCR) und Transaktivierungs-Experimente in Mesophyll-Protoplasten aus Blättern. Wir zeigen, dass MYB112 an eine 8-bp DNA-Fragment, welches die Kernsequenz (A/T/G)(A/C)CC(A/T)(A/G/T)(A/C)(T/C) aufweist. Mit Hilfe von Electrophoretic Mobility Shift Assay (EMSA) und Chromatin-Immunopräzipitation gekoppelt mit qPCR (ChIP-qPCR) zeigen wir, dass MYB112 in vitro und in vivo an die Promotoren von MYB7 und MYB32 bindet was sie damit als direkte Zielgene von MYB112 identifiziert. Es wurde bereits gezeigt, dass MYB TFs eine wichtige Rolle bei der Kontrolle der Flavonoid-Biosynthese in Pflanzen haben. Viele Faktoren, die oberhalb des Anthocyanin-Biosyntheseweges agieren, werden bei Stickstofflimitierung oder erhöhter Saccharose-Konzentration auch verstärkt exprimiert. Außer den erwähnten Bedingungen können auch andere Umweltparameter, wie z. B. erhöhter Salzgehalt und Starklicht zu erhöhter Expression führen. Im Gegensatz jedoch zu einigen anderen MYB TFs, die einen Einfluss auf Gene des Anthocyanin-Biosyntheseweges ausüben, ist die Expression von MYB112 nicht durch Stickstoff-Limitierung oder Kohlenstoffüberfluss kontrolliert, sondern wird durch erhöhten Salzgehalt sowie Starklicht stimuliert. Somit ist MYB112 ein neuer Regulator, der eine Anthocyanin-Akkumulation unter abiotischen Stressbedingungen kontrolliert.

[Truncated abstract] Human complement receptor 2 (CR2/CD21) is a cell surface glycoprotein detected on specific cells involved in immunity, which binds complement C3 cleavage fragments, cellular ligands IFN-? and CD23 as well as the EBV coat protein, gp350/220. During the early stages of B-cell development CR2/CD21 is silenced. Expression is initiated on immature B-cells escaping negative selection. During peripheral maturation CR2/CD21 is up-regulated with B-cell sub-populations showing distinctive surface levels (comparatively low, intermediate or high). CR2/CD21 is silenced upon terminal plasmacytic differentiation. Appropriate timing and expression level of CR2/CD21 is important for the development of a healthy B-cell repertoire. Previous studies have identified sequences within the proximal promoter and first intron of CR2/CD21 that cooperate within native chromatin to control cell-specific silencing. Further, analysis of cultured human cells has revealed chromatin structural variation causing DNase I hypersensitivity at these regulatory sites in a CR2/CD21-expressing mature B-cell line (Raji) which are absent in a non-lymphoid cell type (K562). The primary focus of the present study involved characterising chromatin structural variation over previously recognized DNase I hypersensitive regions at the CR2/CD21 locus in human cells to understand how chromatin structure might regulate developmental expression of CR2/CD21. ... These studies provide evidence that notch signaling influences CR2/CD21 expression in human cell lines. First, in vivo binding of CBF1 to CR2/CD21 sequences in the proximal promoter and CRS implies that CR2/CD21 is a direct target of notch activation. Second, the effect of exogenous notch signalling molecules on CR2/CD21 proximal promoter activity was modulated by factors binding tandem E-boxes near the transcriptional start site suggesting that the notch pathway may also influence CR2/CD21 expression via control of HLH molecules. Third, initiation of CR2/CD21 expression was observed in a nonexpressing pre-B cell line (Reh) by co-culture with stromal cells expressing a notch ligand (OP9-DL) but not control stroma (OP9-GFP). Together, these findings support a role for notch regulation of B-cell maturation and invite speculation that initiation of CR2/CD21 expression following negative selection of immature B-cells involves crosstalk between HLH transcriptional regulators and the notch pathway. Furthermore, the Reh/OP9-DL co-culture system may provide a model to directly study the relationship between cell signalling molecules, transcription factor regulation, chromatin structural variation and differentiation of B-cells.

La chromatine, dont l’unité de base est le nucléosome, est une structure nucléoprotéique dynamique qui nécessite un remodelage au cours des processus nucléaires utilisant l’ADN comme matrice tels que la réplication, la transcription ou la réparation des cassures et autres types de lésions à l’ADN. Plusieurs facteurs capables de moduler la structure de la chromatine ont été caractérisés. Ils regroupent les complexes de remodelage ATP-dépendants et les enzymes modifiant les histones de façon post-traductionnelle. Nous nous intéressons au laboratoire à la voie de méthylation de la lysine 20 située sur la queue aminoterminale de l’histone H4. Le premier niveau de méthylation est induit par la monométhyltransférase PR-Set7 tandis que la di et tri méthylation sont déposées par le couple d’enzymes SUV4-20H1/2. Pour mieux caractériser le rôle joué par PR-Set7 au cours du développement, j’ai étudié la fonction de l'orthologue de PR-Set7 chez la Drosophile (dPR-Set7). La première partie de ma thèse a consisté à caractériser le rôle de dPR-Set7 dans la transcription. De manière intéressante, nous avons montré que la régulation transcriptionnelle médiée par PR-Set7 nécessite son domaine SET enzymatique mais pas H4K20me, suggérant l'existence d’autres substrats non-histones. Nous avons mis en évidence une interaction fonctionnelle de PR-Set7 et ISWI qui est la sous unité catalytique des complexes de remodelage de la chromatine (CRC). Fait intéressant, ISWI contient un patch basique identique à la queue N-terminale de H4 suggérant qu’il pourrait être un substrat pour dPR-Set7. La deuxième partie de ma thèse a consisté à combiner l’effet des radiations à ceux induits par les inhibiteurs des méthyltransférases responsables de la méthylation H4K20 et les radiations dans les lignées de cellules cancéreuses du pancréas et de l’ovaire. Par l'utilisation de différents inhibiteurs ciblant soit PR-Set7 ou les enzymes SUV4-20H, j’ai voulu savoir si la baisse des niveaux de H4K20me pourrait contribuer à une meilleure efficacité des traitements aux rayons X. Nos résultats montrent que la diminution globale des marques H4K20me2/3 suite à l’inhibition des enzymes SUV4-20H n’impacte que faiblement sur la survie des cellules et la combinaison des deux traitements les rendraient plus radiosensibles. / Chromatin is a dynamic nucleoprotein structure that requires remodeling for all nuclear processes such as replication, transcription and DNA damage. Several factors have been characterized to modulate chromatin structure and include ATP-dependent remodeling complexes and histone-modifying enzymes. We are interested in the laboratory in the methylation pathway of lysine 20 on histone H4 tail. The first level of methylation is induced by the monomethyltransferase PR-Set7 and the di/tri methylation are deposited by the SUV4-20H enzymes. To better characterize the role of dPR-Set7 during development, I wanted to study the function of Drosophila PR-Set7 (dPR-Set7). The first part of my thesis aimed to unravel the role of dPR-Set7 in transcription. Interestingly, transcriptional regulation mediated by PR-Set7 requires its enzymatic SET domain but not H4K20me, suggesting the existence of other non-histone substrates. We demonstrated a functional interaction between PR-Set7 and ISWI, the catalytic subunit of chromatin remodeling complexes (CRC). Interestingly, ISWI contains a basic patch identical to the histone H4 tail suggesting that it could be a substrate for dPR-Set7. The second part of my thesis consisted in combining inhibitors of H4K20 methyltransferase and radiation in ovarian and pancreatic cancer cell lines. I wanted to know if the decrease of H4K20me levels by inhibiting either PR-Set7 or SUV4-20H enzymes, contribute to better X-ray treatments. Our results show that the overall decrease of H4K20me2/3 marks following the inhibition of SUV4-20H enzymes has a low impact on cell survival and the combining effects of both treatments sensitize cancer cells.