scholarly journals Hops/Tmub1 Heterozygous Mouse Shows Haploinsufficiency Effect in Influencing p53-Mediated Apoptosis

2021 ◽  
Vol 22 (13) ◽  
pp. 7186
Author(s):  
Simona Ferracchiato ◽  
Nicola Di-Iacovo ◽  
Damiano Scopetti ◽  
Danilo Piobbico ◽  
Marilena Castelli ◽  
...  
Keyword(s):  
Dna Damage ◽  
Tumor Suppressor ◽  
Dna Damage Response ◽  
Stress Responses ◽  
Mammalian Cells ◽  
P53 Protein ◽  
Suppressor Gene ◽  
Mrna Levels ◽  
Damage Response ◽  
Transcript Profiles

HOPS is a ubiquitin-like protein implicated in many aspects of cellular function including the regulation of mitotic activity, proliferation, and cellular stress responses. In this study, we focused on the complex relationship between HOPS and the tumor suppressor p53, investigating both transcriptional and non-transcriptional p53 responses. Here, we demonstrated that Hops heterozygous mice and mouse embryonic fibroblasts exhibit an impaired DNA-damage response to etoposide-induced double-strand breaks when compared to wild-type genes. Specifically, alterations in HOPS levels caused significant defects in the induction of apoptosis, including a reduction in p53 protein level and percentage of apoptotic cells. We also analyzed the effect of reduced HOPS levels on the DNA-damage response by examining the transcript profiles of p53-dependent genes, showing a suggestive deregulation of the mRNA levels for a number of p53-dependent genes. Taken together, these results show an interesting haploinsufficiency effect mediated by Hops monoallelic deletion, which appears to be enough to destabilize the p53 protein and its functions. Finally, these data indicate a novel role for Hops as a tumor-suppressor gene in DNA damage repair in mammalian cells.

scholarly journals Disruption of the Pdcd4 tumor suppressor gene in chicken DT40 cells reveals its role in the DNA-damage response

Oncogene ◽  
2009 ◽  
Vol 28 (42) ◽  
pp. 3758-3764 ◽  
Author(s):  
P Singh ◽  
R Marikkannu ◽  
N Bitomsky ◽  
K -H Klempnauer
Keyword(s):  
Dna Damage ◽  
Tumor Suppressor ◽  
Tumor Suppressor Gene ◽  
Dna Damage Response ◽  
Suppressor Gene ◽  
Damage Response ◽  
Dt40 Cells

scholarly journals Dephosphorylation of the C-terminal Tyrosyl Residue of the DNA Damage-related Histone H2A.X Is Mediated by the Protein Phosphatase Eyes Absent

2009 ◽  
Vol 284 (24) ◽  
pp. 16066-16070 ◽  
Author(s):  
Navasona Krishnan ◽  
Dae Gwin Jeong ◽  
Suk-Kyeong Jung ◽  
Seong Eon Ryu ◽  
Andrew Xiao ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Mammalian Cells ◽  
Protein Tyrosine Phosphatases ◽  
Histone H2a ◽  
Eyes Absent ◽  
Damage Repair ◽  
Damage Response

In mammalian cells, the DNA damage-related histone H2A variant H2A.X is characterized by a C-terminal tyrosyl residue, Tyr-142, which is phosphorylated by an atypical kinase, WSTF. The phosphorylation status of Tyr-142 in H2A.X has been shown to be an important regulator of the DNA damage response by controlling the formation of γH2A.X foci, which are platforms for recruiting molecules involved in DNA damage repair and signaling. In this work, we present evidence to support the identification of the Eyes Absent (EYA) phosphatases, protein-tyrosine phosphatases of the haloacid dehalogenase superfamily, as being responsible for dephosphorylating the C-terminal tyrosyl residue of histone H2A.X. We demonstrate that EYA2 and EYA3 displayed specificity for Tyr-142 of H2A.X in assays in vitro. Suppression of eya3 by RNA interference resulted in elevated basal phosphorylation and inhibited DNA damage-induced dephosphorylation of Tyr-142 of H2A.X in vivo. This study provides the first indication of a physiological substrate for the EYA phosphatases and suggests a novel role for these enzymes in regulation of the DNA damage response.

scholarly journals Overall Cdk activity modulates the DNA damage response in mammalian cells

2009 ◽  
Vol 187 (6) ◽  
pp. 773-780 ◽  
Author(s):  
Antonio Cerqueira ◽  
David Santamaría ◽  
Bárbara Martínez-Pastor ◽  
Miriam Cuadrado ◽  
Oscar Fernández-Capetillo ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Mammalian Cells ◽  
Cell Cycle Progression ◽  
Cyclin Dependent Kinase ◽  
Genome Maintenance ◽  
Cycle Progression ◽  
Damage Response ◽  
Dna Break ◽  
Specific Contribution

In response to DNA damage, cells activate a phosphorylation-based signaling cascade known as the DNA damage response (DDR). One of the main outcomes of DDR activation is inhibition of cyclin-dependent kinase (Cdk) activity to restrain cell cycle progression until lesions are healed. Recent studies have revealed a reverse connection by which Cdk activity modulates processing of DNA break ends and DDR activation. However, the specific contribution of individual Cdks to this process remains poorly understood. To address this issue, we have examined the DDR in murine cells carrying a defined set of Cdks. Our results reveal that genome maintenance programs of postreplicative cells, including DDR, are regulated by the overall level of Cdk activity and not by specific Cdks.

scholarly journals Role of GOLPH3 and TPX2 in Neuroblastoma DNA Damage Response and Cell Resistance to Chemotherapy

2019 ◽  
Vol 20 (19) ◽  
pp. 4764 ◽  
Author(s):  
Marzia Ognibene ◽  
Marina Podestà ◽  
Alberto Garaventa ◽  
Annalisa Pezzolo
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Mammalian Cells ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cells ◽  
Preschool Age ◽  
Human Neuroblastoma Cell ◽  
Human Neuroblastoma ◽  
Therapeutic Benefits ◽  
Damage Response

Neuroblastoma (NB) is an aggressive, relapse-prone infancy tumor of the sympathetic nervous system and is the leading cause of death among preschool age diseases, so the search for novel therapeutic targets is crucial. Golgi phosphoprotein 3 (GOLPH3) has been reported to be involved in the development, and in the DNA damage response, of various human cancers. Golgi dispersal is a common feature of DNA damage response in mammalian cells. Understanding how cells react to DNA damage is essential in order to recognize the systems used to escape from elimination. We induced DNA damage in two human neuroblastoma cell lines by curcumin. The exposure of neuroblastoma cells to curcumin induced: (a) up-regulation of GOLPH3+ cells; (b) augmentation of double-strand breaks; (c) Golgi fragmentation and dispersal throughout the cytoplasm; (d) increase of apoptosis and autophagy; (e) increased expression of TPX2 oncoprotein, able to repair DNA damage. Primary neuroblastoma samples analysis confirmed these observations. Our findings suggest that GOLPH3 expression levels may represent a clinical marker of neuroblastoma patients’ responsiveness to DNA damaging therapies—and of possible resistance to them. Novel molecules able to interfere with GOLPH3 and TPX2 pathways may have therapeutic benefits when used in combination with standard DNA damaging therapeutic agents in neuroblastoma

scholarly journals MORC2 regulates DNA damage response through a PARP1-dependent pathway

2019 ◽  
Vol 47 (16) ◽  
pp. 8502-8520 ◽  
Author(s):  
Lin Zhang ◽  
Da-Qiang Li
Keyword(s):  
Dna Damage ◽  
Zinc Finger ◽  
Chromatin Remodeling ◽  
Dna Damage Response ◽  
Mammalian Cells ◽  
Cellular Response ◽  
Genotoxic Stress ◽  
Underlying Mechanism ◽  
Dna Lesions ◽  
Damage Response

Abstract Microrchidia family CW-type zinc finger 2 (MORC2) is a newly identified chromatin remodeling enzyme with an emerging role in DNA damage response (DDR), but the underlying mechanism remains largely unknown. Here, we show that poly(ADP-ribose) polymerase 1 (PARP1), a key chromatin-associated enzyme responsible for the synthesis of poly(ADP-ribose) (PAR) polymers in mammalian cells, interacts with and PARylates MORC2 at two residues within its conserved CW-type zinc finger domain. Following DNA damage, PARP1 recruits MORC2 to DNA damage sites and catalyzes MORC2 PARylation, which stimulates its ATPase and chromatin remodeling activities. Mutation of PARylation residues in MORC2 results in reduced cell survival after DNA damage. MORC2, in turn, stabilizes PARP1 through enhancing acetyltransferase NAT10-mediated acetylation of PARP1 at lysine 949, which blocks its ubiquitination at the same residue and subsequent degradation by E3 ubiquitin ligase CHFR. Consequently, depletion of MORC2 or expression of an acetylation-defective PARP1 mutant impairs DNA damage-induced PAR production and PAR-dependent recruitment of DNA repair proteins to DNA lesions, leading to enhanced sensitivity to genotoxic stress. Collectively, these findings uncover a previously unrecognized mechanistic link between MORC2 and PARP1 in the regulation of cellular response to DNA damage.

Novel SIRT1 Agonists, SRT501 and SRT2183, Induce Expression of DNA Repair Genes and Apoptosis of Ph− Acute Lymphoblastic Leukemia Cells Alone and in Combination with the HDAC Inhibitor LBH589..

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3083-3083
Author(s):  
Anna Scuto ◽  
Mark Kirschbaum ◽  
Jennifer M Cermak ◽  
Peter Atadja ◽  
Richard Jove
Keyword(s):  
Dna Damage ◽  
Acute Lymphoblastic Leukemia ◽  
Histone Deacetylase ◽  
Dna Damage Response ◽  
Histone Deacetylase Inhibitors ◽  
Lymphoblastic Leukemia ◽  
Philadelphia Chromosome ◽  
Growth Arrest ◽  
Mrna Levels ◽  
Damage Response

Abstract Abstract 3083 Poster Board III-20 Histone Deacetylase Inhibitors (HDACi) such as LBH589, which inhibit the zinc containing catalytic domain of HDAC of classes I, II, and IV, demonstrate activity against various malignancies, particularly lymphoid malignancies. SIRT1 is an NAD+ dependent class III histone deacetylase, which deacetylates histones as well as non-histone proteins and is not affected directly by HDACi such as LBH589. It remains controversial whether inhibition of SIRT1 or its activation is more efficacious in anticancer therapy. We have studied the activity of two novel SIRT1 activators, SRT501 and SRT2183, in Philadelphia chromosome negative acute lymphoblastic leukemia (ALL) cell lines. Both pre B (NALM-6, Reh) and T cell (MOLT-4) ALL lines were treated with either SRT501 or SRT2183, as well as in combination with LBH589 and evaluated for biological and gene expression responses. SRT501 induced growth arrest and apoptosis at doses ranging from 10-100 uM, with even the lowest doses inhibiting growth at 72 hours. SRT2183 is much more potent, with growth arrest and apoptosis induced at doses ranging from 1-20 uM. PCR array analysis revealed that SRT2183 treatment leads to increased mRNA levels of pro-apoptosis, growth arrest, and DNA damage response genes. We have previously demonstrated that the activity of LBH589 is mediated in part through upregulation or acetylation of proteins involved in the DNA damage response pathways. Quantitative real-time PCR confirms that the combination of LBH589 with SRT2183 leads to significantly higher expression of GADD45A and GADD45G than either agent alone. The combination of LBH589 plus SRT2183 showed enhanced inhibition of c-Myc protein levels, phosphorylation of H2A.X, and interestingly, increased acetylation of p53 (acetylation of p53 was not seen with SRT2183 alone). In summary, the novel SIRT1 activators SRT501 and SRT2183 show growth inhibitory and pro-apoptotic activity in Ph- ALL alone and enhanced activity in combination with LBH589. Clinical studies of these agents, particularly in combination with HDACi are warranted. Disclosures Kirschbaum: Novartis: Consultancy. Cermak:Sirtris: Employment. Atadja:Novartis: Employment.

Discovering the Epigenetic Modifications and the Alterations in Cellular DNA Damage Response Pathways That Drive Myelomagenesis; Clinical Implications Into the Biology of Multiple Myeloma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 299-299
Author(s):  
Maria Gkotzamanidou ◽  
Evangelos Terpos ◽  
Petros P. Sfikakis ◽  
Meletios Athanasios Dimopoulos ◽  
Vassilis L. Souliotis
Keyword(s):  
Dna Damage ◽  
Healthy Volunteers ◽  
Dna Damage Response ◽  
Plasma Cells ◽  
P53 Protein ◽  
Chromatin Condensation ◽  
Protein Accumulation ◽  
Damage Response ◽  
Induction Of Apoptosis ◽  
Cellular Dna

Abstract Abstract 299 The aim of this study was to evaluate epigenetic modifications and alterations in cellular DNA damage response pathways that may be implicated in the multistep transformation of myelomagenesis. Peripheral blood mononuclear cells (PBMCs) and plasma cells from bone marrow aspirates were collected from 15 patients with MGUS (8M/7F), 22 with asymptomatic MM (AMM; 10M/12F), 41 patients with symptomatic MM (16M/25F) who underwent autologous stem cell transplantation as part of their first line therapy, and 12 healthy volunteers (7M/5F; only PBMCs). Epigenetics (chromatin condensation, transcription activity) and DNA damage response pathways (melphalan-induced DNA damage formation/repair in four genomic loci including beta-actin, p53, N-ras and delta-globin genes, accumulation of p53 protein and induction of apoptosis) were evaluated. In both PBMCs and plasma cells and in all genomic regions analyzed, significant differences in the local chromatin looseness between the different groups of patients were observed: healthy volunteers<MGUS<AMM<MM (p<0.02 for all comparisons). In PBMCs and plasma cells from all subjects, beta-actin, p53 and N-ras genes were transcriptionally active, while delta-globin gene was silent in all samples from healthy volunteers and MGUS patients. Notably, an induction of the transcription activity of delta-globin gene was found in 10/22 (45.5%) of AMM and 32/41 (78%) of symptomatic MM patients. Following a 5-min treatment of PBMCs with 100μg/ml melphalan or plasma cells with 35μg/ml, the efficiency of DNA damage repair inside all genes analysed was in accordance with that of chromatin condensation and gene expression efficiency at the same genomic loci: healthy volunteers<MGUS<AMM<MM (p<0.04 for all comparisons). In particular, in the N-ras gene, PBMCs from healthy volunteers showed 128.6±38.6 adducts/106 nucleotides, from MGUS patients 114.3±26.7 adducts/106nucleotides, from AMM 96.7±20.9 adducts/106nucleotides, and from symptomatic MM patients 56.6±27.2 adducts/106nucleotides. Similarly, plasma cells from MGUS patients showed 95.7±25.0 adducts/106nucleotides, from AMM patients 70.3±21.9 adducts/106nucleotides, and from symptomatic MM patients 32.3±10.2 adducts/106nucleotides. There was a strong correlation for the DNA damage repair data between PBMCs and plasma cells from the same individuals (R2=0.60, p<0.001). Moreover, following a 5-min exposure of PBMCs and plasma cells with various doses of melphalan (0–120μg/ml), we found that PBMCs from healthy volunteers showed evidence of p53 protein accumulation at melphalan doses as low as 17.9±8.7 μg/ml, from MGUS patients at 29.7±12.5 μg/ml, from AMM patients at 65.6±23.8 μg/ml, and from symptomatic MM patients at 100.2±29.7 μg/ml. Plasma cells from MGUS patients showed evidence of p53 protein accumulation at melphalan doses as low as 20.2±8.9 μg/ml, from AMM patients at 35.2±14.3 μg/ml, while from symptomatic MM patients at 55.3±23.1 μg/ml (p<0.02 for all comparisons). Linear association for the p53 results between PBMCs and plasma cells from the same individuals was observed (R2=0.65, p<0.001). Also, PBMCs and plasma cells were treated with various doses of melphalan (0–120 μg/ml) for 5 min, and the induction of apoptosis was measured 24h later. In accordance with the p53 data, PBMCs from healthy volunteers showed evidence of induction of apoptosis at melphalan doses as low as 13.2±6.9 μg/ml, from MGUS patients at 20.6±10.8 μg/ml, from AMM patients at 51.4±20.3 μg/ml, and from symptomatic MM patients at 89.7±25.1 μg/ml. Plasma cells from MGUS patients showed induction of apoptosis at melphalan doses as low as 9.9±2.9 μg/ml, from AMM patients at 25.3±8.5 μg/ml, and from symptomatic MM patients at 45.2±19.4 μg/ml (p<0.01 for all comparisons). For apoptosis data, a strong correlation was also found between PBMCs and plasma cells from the same individuals (R2=0.61, p<0.001). In conclusion, our data indicate that myelomagenesis is associated with epigenetic alterations and modifications in the cellular DNA damage response pathways that can be used as novel molecular biomarkers for early diagnosis and prediction of clinical outcome in MM. Furthermore, in all end-points examined, a strong association between PBMCs and plasma cells from the same individuals was observed, suggesting that measurement of these novel molecular biomarkers can be performed in a readily accessible tissue such as PBMCs. Disclosures: No relevant conflicts of interest to declare.

scholarly journals PML Colocalizes with and Stabilizes the DNA Damage Response Protein TopBP1

2003 ◽  
Vol 23 (12) ◽  
pp. 4247-4256 ◽  
Author(s):  
Zhi-Xiang Xu ◽  
Anna Timanova-Atanasova ◽  
Rui-Xun Zhao ◽  
Kun-Sang Chang
Keyword(s):  
Dna Damage ◽  
Dna Damage Response ◽  
Genome Stability ◽  
Growth Regulation ◽  
Suppressor Gene ◽  
Nuclear Body ◽  
Promyelocytic Leukemia ◽  
Multifunctional Protein ◽  
Damage Response

ABSTRACT The PML tumor suppressor gene is consistently disrupted by t(15;17) in patients with acute promyelocytic leukemia. Promyelocytic leukemia protein (PML) is a multifunctional protein that plays essential roles in cell growth regulation, apoptosis, transcriptional regulation, and genome stability. Our study here shows that PML colocalizes and associates in vivo with the DNA damage response protein TopBP1 in response to ionizing radiation (IR). Both PML and TopBP1 colocalized with the IR-induced bromodeoxyuridine single-stranded DNA foci. PML and TopBP1 also colocalized with Rad50, Brca1, ATM, Rad9, and BLM. IR and interferon (IFN) coinduce the expression levels of both TopBP1 and PML. In PML-deficient NB4 cells, TopBP1 was unable to form IR-induced foci. All-trans-retinoic acid induced reorganization of the PML nuclear body (NB) and reappearance of the IR-induced TopBP1 foci. Inhibition of PML expression by siRNA is associated with a significant decreased in TopBP1 expression. Furthermore, PML-deficient cells express a low level of TopBP1, and its expression cannot be induced by IR or IFN. Adenovirus-mediated overexpression of PML in PML−/− mouse embryo fibroblasts substantially increased TopBP1 expression, which colocalized with the PML NBs. These studies demonstrated a mechanism of PML-dependent expression of TopBP1. PML overexpression induced TopBP1 protein but not the mRNA expression. Pulse-chase labeling analysis demonstrated that PML overexpression stabilized the TopBP1 protein, suggesting that PML plays a role in regulating the stability of TopBP1 in response to IR. Together, our findings demonstrate that PML regulates TopBP1 functions by association and stabilization of the protein in response to IR-induced DNA damage.

scholarly journals Normal ABL1 Is a Tumor Suppressor and Therapeutic Target In BCR-ABL1–positive Leukemias

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1466-1466
Author(s):  
Yashodhara Dasgupta ◽  
Mateusz Koptyra ◽  
Margaret Nieborowska-Skorska ◽  
Elisabeth Bolton Gillespie ◽  
Tomasz Stoklosa ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dna Damage ◽  
Tumor Suppressor ◽  
Dna Damage Response ◽  
Chromosome 9 ◽  
Myeloid Differentiation ◽  
Damage Response ◽  
Kinase Expression ◽  
Stimulation Of

Abstract BCR-ABL1 results from t(9;22)(q34;q11) reciprocal translocation resulting in BCR-ABL1 kinase expression, initiating chronic myeloid leukemia in chronic phase (CML-CP). At the initial stages of CML-CP both oncogenic BCR-ABL1 kinase and normal ABL1 kinase are expressed, however, loss of ABL1 kinase expression in CML-CP can result from an interstitial deletion in the normal chromosome 9 [del(9q34)] which may be combined with the transcriptional silencing of the alternative ABL1 promoter within the translocation eventually leading to disease progression and drug resistance. We found that BCR-ABL1 Abl1-/- cells generated a CML-blast phase (BP)-like disease phenotype in SCID mice compared to CML-CP-like disease from BCR-ABL1 Abl1+/+ cells. To determine the mechanisms responsible for blastic transformation of BCR-ABL1 Abl1-/- cells, we examined the role of ABL1 in proliferation, differentiation, apoptosis, genomic instability, and stemness. The presence of ABL1 inhibited proliferation in BCR-ABL1 cells as BCR-ABL1 Abl1-/- cells had higher clonogenic activity and proliferative rate compared to their wild-type counterparts. ABL1 is essential for myeloid differentiation since BCR-ABL1 Abl1-/- cells showed an immature blast phenotype when stained with Wright-Giemsa and myeloid differentiation markers Gr-1 and CD11b. ABL1 promoted apoptosis in response to genotoxic stress as revealed by reduced clonogenicity and elevated expression of p53, phosphoserine-15 p53 and activated caspase 3 in BCR-ABL1 Abl1 +/+ compared to knock-out cells. Although the absence of ABL1 did not enhance ROS and oxidative DNA damage, it appears that an impaired DNA damage response may be responsible for higher chromosome numbers and an accumulation of high numbers of chromosomal aberrations in BCR-ABL1 Abl1-/- cells. We detected an expansion of Lin-c-Kit+Sca-1+ leukemia stem cells (LSCs) in BCR-ABL1 Abl1-/- cells compared to BCR-ABL1 Abl1+/+ or non-transformed counterparts; among the LSCs, there was a higher percentage of CD34-Flt3- long-term and CD34+Flt3-short-term stem cells. These results showed that ABL1 is involved in regulating the LSC compartment in BCR-ABL1 cells. DNA microarray analysis revealed changes in mRNA levels of several genes involved in proliferation, myeloid differentiation, apoptosis, DNA damage response and stemness in BCR-ABL1 Abl1-/- cells in comparison to BCR-ABL1 Abl1+/+ cells. Together, these results demonstrated a critical role of ABL1 in BCR-ABL1-induced leukemia, prolonging survival in mice by suppressing proliferation and expansion of LSC, inducing myeloid differentiation, apoptosis and DNA damage response in BCR-ABL1 cells. Thus, it appears that ABL1 acts as a tumor suppressor in BCR-ABL1 –positive CML cells. Moreover, we hypothesized that the enhancement of the tumor suppressor function of ABL1 may have a significant impact on CML treatment. A small molecule activator of ABL1 kinase, 5-(1,3-diaryl-1H-pyrazol-4-yl)hydantoin (DPH), had been reported to interact with the myristoyl-binding site of ABL1 and destabilize the bent conformation of the α-1 helix, thereby preventing the auto-inhibitory conformation. DPH partially restored ABL1 activity in imatinib-treated cells. DPH-mediated stimulation of ABL1 tumor suppressor activity enhanced the effect of imatinib and ponatinib against CML CD34+ cells, Philadelphia chromosome-positive B-ALL (Ph+B-ALL) cells and relapsed Ph+B-ALL cells harboring T315I mutation without affecting normal counterparts. In summary, ABL1 is a potential tumor suppressor in BCR-ABL1-induced leukemia and stimulation of its function may play a significant role in the development of novel therapeutic strategies for CML and Ph+ALL. Disclosures: No relevant conflicts of interest to declare.

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