Jeroen Essers

Erasmus MC, Rotterdam, South Holland, Netherlands

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Publications (76)560.25 Total impact

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    ABSTRACT: Genome maintenance by homologous recombination depends on coordinating many proteins in time and space to assemble at DNA break sites. To understand this process, we followed the mobility of BRCA2, a critical recombination mediator, in live cells at the single-molecule level using both single-particle tracking and fluorescence correlation spectroscopy. BRCA2-GFP and -YFP were compared to distinguish diffusion from fluorophore behavior. Diffusive behavior of fluorescent RAD51 and RAD54 was determined for comparison. All fluorescent proteins were expressed from endogenous loci. We found that nuclear BRCA2 existed in oligomeric clusters, and exhibited heterogeneous mobility. DNA damage increased BRCA2 transient binding, presumably including binding to damaged sites. Despite its very different size, RAD51 displayed mobility similar to BRCA2, which indicates physical interaction between these proteins both before and after induction of DNA damage. We propose that BRCA2-mediated sequestration of nuclear RAD51 serves to prevent inappropriate DNA interactions and that all RAD51 is delivered to DNA damage sites in association with BRCA2. © 2014 Reuter et al.
    The Journal of cell biology. 12/2014; 207(5):599-613.
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    ABSTRACT: Accurate DNA double-strand break repair through homologous recombination is essential for preserving genome integrity. Disruption of the gene encoding RAD51, the protein that catalyzes DNA strand exchange during homologous recombination, results in lethality of mammalian cells. Proteins required for homologous recombination, also play an important role during DNA replication. To explore the role of RAD51 in DNA replication and DSB repair, we used a knock-in strategy to express a carboxy-terminal fusion of green fluorescent protein to mouse RAD51 (mRAD51-GFP) in mouse embryonic stem cells. Compared to wild-type cells, heterozygous mRad51(+/wt-GFP) embryonic stem cells showed increased sensitivity to DNA damage induced by ionizing radiation and mitomycin C. Moreover, gene targeting was found to be severely impaired in mRad51(+/wt-GFP) embryonic stem cells. Furthermore, we found that mRAD51-GFP foci were not stably associated with chromatin. From these experiments we conclude that this mRad51-GFP allele is an antimorphic allele. When this allele is present in a heterozygous condition over wild-type mRad51, embryonic stem cells are proficient in DNA replication but display defects in homologous recombination and DNA damage repair. Copyright © 2014 Elsevier B.V. All rights reserved.
    DNA repair. 11/2014; 25C:27-40.
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    ABSTRACT: A side effect of radiation therapy in the head and neck region is injury to surrounding healthy tissues such as irreversible impaired function of the salivary glands. Hyperbaric oxygen therapy (HBOT) is clinically used to treat radiation-induced damage but its mechanism of action is largely unknown. In this study, we investigated the molecular pathways that are affected by HBOT in mouse salivary glands two weeks after radiation therapy by microarray analysis. Interestingly, HBOT led to significant attenuation of the radiation induced expression of a set of genes and upstream regulators that are involved in processes like fibrosis and tissue regeneration. Our data suggest that the TGFβ-pathway, which is involved in radiation-induced fibrosis and chronic loss of function after radiation therapy, is affected by HBOT. On the longer term, HBOT reduced the expression of the fibrosis-associated factor α-smooth muscle actin in irradiated salivary glands. This study highlights the potential of HBOT to inhibit the TGFβ-pathway in irradiated salivary glands and restrain consequential radiation induced tissue injury.
    Molecular medicine (Cambridge, Mass.). 05/2014;
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    ABSTRACT: Loeys-Dietz syndrome (LDS) is an autosomal dominant arterial aneurysm disease belonging to the spectrum of transforming growth factor β (TGFβ)-associated vasculopathies. In its most typical form it is characterized by the presence of hypertelorism, bifid uvula/cleft palate and aortic aneurysm and/or arterial tortuosity. LDS is caused by heterozygous loss of function mutations in the genes encoding TGFβ receptor 1 and 2 (TGFBR1 and -2), which lead to a paradoxical increase in TGFβ signaling. To address this apparent paradox and to gain more insight into the pathophysiology of aneurysmal disease, we characterized a new Tgfbr1 mouse model carrying a p.Y378* nonsense mutation. Study of the natural history in this model showed that homozygous mutant mice die during embryonic development due to defective vascularization. Heterozygous mutant mice aged 6 and 12 months were morphologically and (immuno)histochemically indistinguishable from wild-type mice. We show that the mutant allele is degraded by nonsense mediated mRNA decay, expected to result in haploinsufficiency of the mutant allele. Since this haploinsufficiency model does not result in cardiovascular malformations, it does not allow further study of the process of aneurysm formation. In addition to providing a comprehensive method for cardiovascular phenotyping in mice, the results of this study confirm that haploinsuffciency is not the underlying genetic mechanism in human LDS.
    PLoS ONE 01/2014; 9(2):e89749. · 3.53 Impact Factor
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    ABSTRACT: In this study we set out to investigate the clinically observed relationship between chronic obstructive pulmonary disease (COPD) and aortic aneurysms. We tested the hypothesis that an inherited deficiency of connective tissue might play a role in the combined development of pulmonary emphysema and vascular disease.
    PLoS ONE 01/2014; 9(9):e106054. · 3.53 Impact Factor
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    ABSTRACT: Caffeine is a widely used inhibitor of the protein kinases that play a central role in the DNA damage response. We used chemical inhibitors and genetically deficient mouse embryonic stem cell lines to study the role of DNA damage response in stable integration of the transfected DNA and found that caffeine rapidly, efficiently and reversibly inhibited homologous integration of the transfected DNA as measured by several homologous recombination-mediated gene-targeting assays. Biochemical and structural biology experiments revealed that caffeine interfered with a pivotal step in homologous recombination, homologous joint molecule formation, through increasing interactions of the RAD51 nucleoprotein filament with non-homologous DNA. Our results suggest that recombination pathways dependent on extensive homology search are caffeine-sensitive and stress the importance of considering direct checkpoint-independent mechanisms in the interpretation of the effects of caffeine on DNA repair.
    Nucleic Acids Research 05/2013; · 8.81 Impact Factor
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    ABSTRACT: DNA double-strand breaks (DSBs) occur frequently during replication in sister chromatids, and are dramatically increased when cells are exposed to chemotherapeutic agents including camptothecin. Such DSBs are efficiently repaired specifically by homologous recombination (HR) with the intact sister chromatid. HR hence plays pivotal roles in cellular proliferation and cellular tolerance to camptothecin. Mammalian cells carry several structure-specific endonucleases, such as Xpf-Ercc1 and Mus81-Eme1, in which Xpf and Mus81 are the essential subunits for enzymatic activity. Here we show the functional overlap between Xpf and Mus81 by conditionally inactivating Xpf in the chicken DT40 cell line, which has no Mus81 ortholog. Although mammalian cells deficient in either Xpf or Mus81 are viable, Xpf inactivation in DT40 cells was lethal, resulting in a marked increase in the number of spontaneous chromosome breaks. Similarly, inactivation of both Xpf and Mus81 in human HeLa cells and murine embryonic stem cells caused numerous spontaneous chromosome breaks. Furthermore, the phenotype of Xpf-deficient DT40 cells was reversed by ectopic expression of human Mus81-Eme1 or human Xpf-Ercc1 heterodimers. These observations indicate the functional overlap of Xpf-Ercc1 and Mus81-Eme1 in the maintenance of genomic DNA. Both Mus81-Eme1 and Xpf-Ercc1 contribute to the completion of HR as evidenced by the following data that the expression of Mus81-Eme1 or Xpf-Ercc1 diminished the number of camptothecin-induced chromosome breaks in Xpf-deficient DT40 cells, and preventing early steps in HR by deleting XRCC3 suppressed the inviability of Xpf-deficient DT40 cells. In summary, Xpf and Mus81 have a substantially overlapping function in completion of HR.
    Cancer Research 04/2013; · 9.28 Impact Factor
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    ABSTRACT: The ten-subunit transcription factor IIH (TFIIH) plays a crucial role in transcription and nucleotide excision repair (NER). Inactivating mutations in the smallest 8-kDa TFB5/TTDA subunit cause the neurodevelopmental progeroid repair syndrome trichothiodystrophy A (TTD-A). Previous studies have shown that TTDA is the only TFIIH subunit that appears not to be essential for NER, transcription, or viability. We studied the consequences of TTDA inactivation by generating a Ttda knock-out (Ttda(-/-) ) mouse-model resembling TTD-A patients. Unexpectedly, Ttda(-/-) mice were embryonic lethal. However, in contrast to full disruption of all other TFIIH subunits, viability of Ttda(-/-) cells was not affected. Surprisingly, Ttda(-/-) cells were completely NER deficient, contrary to the incomplete NER deficiency of TTD-A patient-derived cells. We further showed that TTD-A patient mutations only partially inactivate TTDA function, explaining the relatively mild repair phenotype of TTD-A cells. Moreover, Ttda(-/-) cells were also highly sensitive to oxidizing agents. These findings reveal an essential role of TTDA for life, nucleotide excision repair, and oxidative DNA damage repair and identify Ttda(-/-) cells as a unique class of TFIIH mutants.
    PLoS Genetics 04/2013; 9(4):e1003431. · 8.52 Impact Factor
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    ABSTRACT: Rad54 is an important factor in the homologous recombination pathway of DNA double-strand break repair. However, Rad54 knockout (KO) mice do not exhibit overt phenotypes at adulthood, even when exposed to radiation. In this study, we show that in Rad54 KO mouse the germline is actually altered. Compared with the wild-type (WT) animals, these mice have less premeiotic germ cells. This germ cell loss is found as early as in E11.5 embryos, suggesting an early failure during mutant primordial germ cells development. Both testicular and ovarian KO germ cells exhibited high radiation sensitivity leading to a long-term gametogenesis defect at adulthood. The KO female germline was particularly affected displaying decreased litter size or sterility. Spermatogenesis recovery after irradiation was slower and incomplete in Rad54 KO mice compared with that of WT mice, suggesting that loss of germ stem cell precursors is not fully compensated along the successive rounds of spermatogenesis. Finally, spermatogenesis recovery after postnatal irradiation is in part regulated by glial-cell-line-derived neurotrophic factor (GDNF) in KO but not in irradiated WT mice, suggesting that Sertoli cell GDNF production is stimulated upon substantial germ cell loss only. Our findings suggest that Rad54 has a key function in maintaining genomic integrity of the developing germ cells.
    Cell Death & Disease 01/2013; 4:e774. · 6.04 Impact Factor
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    ABSTRACT: In S and G2 phase mammalian cells DNA double strand breaks (DSBs) can potentially be repaired by homologous recombination (HR) or non-homologous end-joining (NHEJ). Results of several studies suggest that these two mechanistically distinct repair pathways can compete for DNA ends. Because HR and NHEJ differ with respect to error susceptibility, generation of chromosome rearrangements, which are potentially carcinogenic products of DSB repair, may depend on the pathway choice. To investigate this hypothesis, the influence of HR and NHEJ inhibition on the frequencies of chromosome aberrations in G2 phase cells was investigated. SW-1573 and RKO cells were treated with mild (41°C) hyperthermia in order to disable HR and/or NU7441/cisplatin to inactivate NHEJ and frequencies of chromosomal fragments (resulting from unrepaired DSBs) and translocations (products of erroneous DSB rejoining) were studied using premature chromosome condensation (PCC) combined with fluorescence in situ hybridization (FISH). It is shown here that temporary inhibition of HR by hyperthermia results in increased frequency of ionizing-radiation (IR)-induced chromosomal translocations and that this effect is abrogated by NU7441- or cisplatin-mediated inhibition of NHEJ. The results suggest that in the absence of HR, DSB repair is shifted to the error-prone NHEJ pathway resulting in increased frequencies of chromosomal rearrangements. These results might be of consequence for clinical cancer treatment approaches that aim at inhibition of one or more DSB repair pathways.
    DNA repair 12/2012; · 4.20 Impact Factor
  • ASME 2012 Summer Bioengineering Conference, Parts A and B; 06/2012
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    ABSTRACT: Telomere maintenance and DNA repair are important processes that protect the genome against instability. mRtel1, an essential helicase, is a dominant factor setting telomere length in mice. In addition, mRtel1 is involved in DNA double-strand break repair. The role of mRtel1 in telomere maintenance and genome stability is poorly understood. Therefore we used mRtel1-deficient mouse embryonic stem cells to examine the function of mRtel1 in replication, DNA repair, recombination, and telomere maintenance. mRtel1-deficient mouse embryonic stem cells showed sensitivity to a range of DNA-damaging agents, highlighting its role in replication and genome maintenance. Deletion of mRtel1 increased the frequency of sister chromatid exchange events and suppressed gene replacement, demonstrating the involvement of the protein in homologous recombination. mRtel1 localized transiently at telomeres and is needed for efficient telomere replication. Of interest, in the absence of mRtel1, telomeres in embryonic stem cells appeared relatively stable in length, suggesting that mRtel1 is required to allow extension by telomerase. We propose that mRtel1 is a key protein for DNA replication, recombination, and repair and efficient elongation of telomeres by telomerase.
    Molecular biology of the cell 05/2012; 23(14):2782-92. · 5.98 Impact Factor
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    ABSTRACT: We characterized the in vivo importance of the homologous recombination factor RAD54 for the developing mouse brain cortex in normal conditions or after ionizing radiation exposure. Contrary to numerous homologous recombination genes, Rad54 disruption did not impact the cortical development without exogenous stress, but it dramatically enhanced the radiation sensitivity of neural stem and progenitor cells. This resulted in the death of all cells irradiated during S or G2, whereas the viability of cells irradiated in G1 or G0 was not affected by Rad54 disruption. Apoptosis occurred after long arrests at intra-S and G2/M checkpoints. This concerned every type of neural stem and progenitor cells, showing that the importance of Rad54 for radiation response was linked to the cell cycle phase at the time of irradiation and not to the differentiation state. In the developing brain, RAD54-dependent homologous recombination appeared absolutely required for the repair of damages induced by ionizing radiation during S and G2 phases, but not for the repair of endogenous damages in normal conditions. Altogether our data support the existence of RAD54-dependent and -independent homologous recombination pathways.
    PLoS ONE 01/2012; 7(5):e37194. · 3.53 Impact Factor
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    ABSTRACT: The DNA damage response (DDR) is a complex signaling network that leads to damage repair while modulating numerous cellular processes. DNA double-strand breaks (DSBs), a highly cytotoxic DNA lesion, activate this system most vigorously. The DSB response network is orchestrated by the ATM protein kinase, which phosphorylates key players in its various branches. Proteasome-mediated protein degradation plays an important role in the proteome dynamics following DNA damage induction. Here, we identify the nuclear proteasome activator PA28γ (REGγ; PSME3) as a novel DDR player. PA28γ depletion leads to cellular radiomimetic sensitivity and a marked delay in DSB repair. Specifically, PA28γ deficiency abrogates the balance between the two major DSB repair pathways--nonhomologous end-joining and homologous recombination repair. Furthermore, PA28γ is found to be an ATM target, being recruited to the DNA damage sites and required for rapid accumulation of proteasomes at these sites. Our data reveal a novel ATM-PA28γ-proteasome axis of the DDR that is required for timely coordination of DSB repair.
    Cell cycle (Georgetown, Tex.) 12/2011; 10(24):4300-10. · 5.24 Impact Factor
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    ABSTRACT: Aging represents a major risk factor for coronary artery disease and aortic aneurysm formation. MicroRNAs (miRs) have emerged as key regulators of biological processes, but their role in age-associated vascular pathologies is unknown. We aim to identify miRs in the vasculature that are regulated by age and play a role in age-induced vascular pathologies. Expression profiling of aortic tissue of young versus old mice identified several age-associated miRs. Among the significantly regulated miRs, the increased expression of miR-29 family members was associated with a profound downregulation of numerous extracellular matrix (ECM) components in aortas of aged mice, suggesting that this miR family contributes to ECM loss, thereby sensitizing the aorta for aneurysm formation. Indeed, miR-29 expression was significantly induced in 2 experimental models for aortic dilation: angiotensin II-treated aged mice and genetically induced aneurysms in Fibulin-4(R/R) mice. More importantly, miR-29b levels were profoundly increased in biopsies of human thoracic aneurysms, obtained from patients with either bicuspid (n=79) or tricuspid aortic valves (n=30). Finally, LNA-modified antisense oligonucleotide-mediated silencing of miR-29 induced ECM expression and inhibited angiotensin II-induced dilation of the aorta in mice. In conclusion, miR-29-mediated downregulation of ECM proteins may sensitize the aorta to the formation of aneurysms in advanced age. Inhibition of miR-29 in vivo abrogates aortic dilation in mice, suggesting that miR-29 may represent a novel molecular target to augment matrix synthesis and maintain vascular wall structural integrity.
    Circulation Research 09/2011; 109(10):1115-9. · 11.86 Impact Factor
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    ABSTRACT: Ultraviolet (UV) radiation-induced DNA lesions can be efficiently repaired by nucleotide excision repair (NER). However, NER is less effective during replication of UV-damaged chromosomes. In contrast, translesion DNA synthesis (TLS) and homologous recombination (HR) are capable of dealing with lesions in replicating DNA. The core HR protein in mammalian cells is the strand exchange protein RAD51, which is aided by numerous proteins, including RAD54. We used RAD54 as a cellular marker for HR to study the response of mammalian embryonic stem (ES) cells to UV irradiation. In contrast to yeast, ES cells lacking RAD54 are not UV sensitive. Here we show that the requirement for mammalian RAD54 is masked by active NER. By genetically inactivating NER and HR through disruption of the Xpa and Rad54 genes, respectively, we demonstrate the contribution of HR to chromosomal integrity upon UV irradiation. We demonstrate using chromosome fiber analysis at the individual replication fork level, that HR activity is important for the restart of DNA replication after induction of DNA damage by UV-light in NER-deficient cells. Furthermore, our data reveal RAD54-dependent and -independent contributions of HR to the cellular sensitivity to UV-light, and they uncover that RAD54 can compensate for the loss of TLS polymerase η with regard to UV-light sensitivity. In conclusion, we show that HR is important for the progression of UV-stalled replication forks in ES cells, and that protection of the fork is an interplay between HR and TLS.
    DNA repair 08/2011; 10(11):1095-105. · 4.20 Impact Factor
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    ABSTRACT: Thoracic aortic aneurysms (TAAs) are a potential life-threatening disease with limited pharmacological treatment options. Current treatment options are aimed at lowering aortic hemodynamic stress, predominantly with β-adrenoceptor blockers. Increasing evidence supports a role for the renin-angiotensin system (RAS) in aneurysm development. RAS blockade would not only lower blood pressure, but might also target the molecular pathways involved in aneurysm formation, in particular the transforming growth factor-β and extracellular signal-regulated kinase 1/2 pathways. Indeed, the angiotensin II type 1 (AT₁) receptor blocker losartan was effective in lowering aortic root growth in mice and patients with Marfan's syndrome. RAS inhibition (currently possible at 3 levels, i.e. renin, ACE and the AT₁ receptor) is always accompanied by a rise in renin due to interference with the negative feedback loop between renin and angiotensin II. Only during AT₁ receptor blockade will this result in stimulation of the non-blocked angiotensin II type 2 (AT₂) receptor. This review summarizes the clinical aspects of TAAs, provides an overview of the current mouse models for TAAs, and focuses on the RAS as a new target for TAA treatment, discussing in particular the possibility that AT₂ receptor stimulation might be crucial in this regard. If true, this would imply that AT₁ receptor blockers (and not ACE inhibitors or renin inhibitors) should be the preferred treatment option for TAAs.
    Pharmacology [?] Therapeutics 07/2011; 131(1):50-60. · 7.79 Impact Factor
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    ABSTRACT: Defective homologous recombination (HR) DNA repair imposed by BRCA1 or BRCA2 deficiency sensitizes cells to poly (ADP-ribose) polymerase (PARP)-1 inhibition and is currently exploited in clinical treatment of HR-deficient tumors. Here we show that mild hyperthermia (41-42.5 °C) induces degradation of BRCA2 and inhibits HR. We demonstrate that hyperthermia can be used to sensitize innately HR-proficient tumor cells to PARP-1 inhibitors and that this effect can be enhanced by heat shock protein inhibition. Our results, obtained from cell lines and in vivo tumor models, enable the design of unique therapeutic strategies involving localized on-demand induction of HR deficiency, an approach that we term induced synthetic lethality.
    Proceedings of the National Academy of Sciences 06/2011; 108(24):9851-6. · 9.81 Impact Factor
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    ABSTRACT: Heterozygous Patched1 (Ptc1(+/-)) mice are prone to medulloblastoma (MB), and exposure of newborn mice to ionizing radiation dramatically increases the frequency and shortens the latency of MB. In Ptc1(+/-) mice, MB is characterized by loss of the normal remaining Ptc1 allele, suggesting that genome rearrangements may be key events in MB development. Recent evidence indicates that brain tumors may be linked to defects in DNA-damage repair processes, as various combinations of targeted deletions in genes controlling cell-cycle checkpoints, apoptosis and DNA repair result in MB in mice. Non-homologous end joining (NHEJ) and homologous recombination (HR) contribute to genome stability, and deficiencies in either pathway predispose to genome rearrangements. To test the role of defective HR or NHEJ in tumorigenesis, control and irradiated Ptc1(+/-) mice with two, one or no functional Rad54 or DNA-protein kinase catalytic subunit (DNA-PKcs) alleles were monitored for MB development. We also examined the effect of Rad54 or DNA-PKcs deletion on the processing of endogenous and radiation-induced double-strand breaks (DSBs) in neural precursors of the developing cerebellum, the cells of origin of MB. We found that, although HR and NHEJ collaborate in protecting cells from DNA damage and apoptosis, they have opposite roles in MB tumorigenesis. In fact, although Rad54 deficiency increased both spontaneous and radiation-induced MB development, DNA-PKcs disruption suppressed MB tumorigenesis. Together, our data provide the first evidence that Rad54-mediated HR in vivo is important for suppressing tumorigenesis by maintaining genomic stability.
    Oncogene 05/2011; 30(47):4740-9. · 8.56 Impact Factor

Publication Stats

3k Citations
560.25 Total Impact Points

Institutions

  • 2004–2013
    • Erasmus MC
      • • Department of Genetics
      • • Department of Oncological Surgery
      Rotterdam, South Holland, Netherlands
    • University of Amsterdam
      • Faculty of Medicine AMC
      Amsterdam, North Holland, Netherlands
  • 2010
    • Leiden University Medical Centre
      Leyden, South Holland, Netherlands
  • 2000–2007
    • Max Planck Institute of Molecular Cell Biology and Genetics
      Dresden, Saxony, Germany
  • 1997–2007
    • Erasmus Universiteit Rotterdam
      • Department of Pathology
      Rotterdam, South Holland, Netherlands