Interaction between ATM and PARP-1 in response to DNA damage and sensitization of ATM deficient cells through PARP inhibition

ATM and PARP-1 are two of the most important players in the cell's response to DNA damage. PARP-1 and ATM recognize and bound to both single and double strand DNA breaks in response to different triggers. Here we report that ATM and PARP-1 form a molecular complex in vivo in undamaged cells and this association increases after γ-irradiation. ATM is also modified by PARP-1 during DNA damage. We have also evaluated the impact of PARP-1 absence or inhibition on ATM-kinase activity and have found that while PARP-1 deficient cells display a defective ATM-kinase activity and reduced γ-H2AX foci formation in response to γ-irradiation, PARP inhibition on itself is able to activate ATM-kinase. PARP inhibition induced γ H2AX foci accumulation, in an ATM-dependent manner. Inhibition of PARP also induces DNA double strand breaks which were dependent on the presence of ATM. As consequence ATM deficient cells display an increased sensitivity to PARP inhibition. In summary our results show that while PARP-1 is needed in the response of ATM to gamma irradiation, the inhibition of PARP induces DNA double strand breaks (which are resolved in and ATM-dependent pathway) and activates ATM kinase

Clinical and Cytokine Profile in Patients with Early and Late Onset Meniere Disease

Background: Meniere disease (MD) is an inner ear disorder associated with comorbidities such as autoimmune diseases or migraine. This study describes clinical and cytokine profiles in MD according to the age of onset of the condition. Methods: A cross-sectional study including 83 MD patients: 44 with early-onset MD (EOMD, <35 years old), and 39 with late-onset MD (LOMD, >50 years old), 64 patients with migraine and 55 controls was carried out. Clinical variables and cytokines levels of CCL3, CCL4, CCL18, CCL22, CXCL,1 and IL-1β were compared among the different groups. Results: CCL18 levels were higher in patients with migraine or MD than in controls. Elevated levels of IL-1β were observed in 11.4% EOMD and in 10.3% LOMD patients and these levels were not dependent on the age of individuals. EOMD had a longer duration of the disease (p = 0.004) and a higher prevalence of migraine than LOMD (p = 0.045). Conclusions: Patients with EOMD have a higher prevalence of migraine than LOMD, but migraine is not associated with any cytokine profile in patients with MD. The levels of CCL18, CCL3, and CXCL4 were different between patients with MD or migraine and controls.ISCIII and European Regional Funds (Grants PI17/01644 and PI20/01126)Andalusian Health Government (Grant PE-0356-2018).Andalusian Health Government (Grant PI-0027-2020)

Efecto de la inhibición de PARP-1 sobre angiogénesis, metástasis y mimetismo vasculogénico en melanoma

La inhibición de PARP pueden inducir efectos anti tumorales cuando se usa como monoterapia o en combinación con la quimioterapia o la radioterapia en tumores diversos ajustes, sin embargo, la base para las actividades contra la metástasis como resultado de la inhibición de PARP sigue siendo una incógnita. El análisis proteómico de las células endoteliales reveló que la vimentina, un filamento de intermediarios involucrados en la angiogénesis y el marcador específico del EndMT (transición endotelio mesénquimal), disminuye bajo la pérdida de la función de PARP-1 en las células endoteliales. Asimismo, en este trabajo se observó que la VE-cadherina, un marcador endotelial vascular de la normalización, se reguló en HUVEC bajo tratamientos con inhibidores de PARP o el silenciamiento de PARP-1. En las células del melanoma, la inhibición de PARP fué capaz de reducir marcadores pro-angiogénicos, tanto en cultivos celulares como en modelos in vivo Hemos demostrado que la vimetina por sí sola es suficiente para inducir el aumento de los efectos fenotípicos mesenquimales / prometastasicos en las células del melanoma, incluyendo OLK/GSK 3¿b dependiente E-cadherina bajo regulada, Snail 1 junto a la activación y la motilidad celular y el aumentó de la migración. En un modelo murino de melanoma metastásico, la inhibición de PARP puede contrarrestar la capacidad de las células del melanoma para formar metástasis en el pulmón. Los ratones implantados con células de melanoma metastático, que fueron tratados también con el inhibidor de PARP DPQ, exhibieron una drástica disminución tanto en la metástasis de pulmón como en la angiogénesis tumoral en comparación con el grupo de ratones control. El grupo de ratones tratados también experimentó un aumento de la supervivencia en comparación con el grupo control. Un silenciamiento estable de PARP-1 en células de melanoma también dio lugar a un aumento significativo en la supervivencia. Estos resultados sugieren que la inhibición de PARP interfiere con los procesos clave que promueven la metástasis, como el cambio en el fenotipo que permite a las células de melanoma adquirir propiedades invasivas.Tesis Univ. Granada. Departamento de Bioquímica y Biología Molecular III e Inmunologí

ROS-induced DNA damage and PARP-1 are required for optimal induction of starvation-induced autophagy

Rodríguez-Vargas, José Manuel et al.In response to nutrient stress, cells start an autophagy program that can lead to adaptation or death. The mechanisms underlying the signaling from starvation to the initiation of autophagy are not fully understood. In the current study we show that the absence or inactivation of PARP-1 strongly delays starvation-induced autophagy. We have found that DNA damage is an early event of starvation-induced autophagy as measured by ¿-H2AX accumulation and comet assay, with PARP-1 knockout cells displaying a reduction in both parameters. During starvation, ROS-induced DNA damage activates PARP-1, leading to ATP depletion (an early event after nutrient deprivation). The absence of PARP-1 blunted AMPK activation and prevented the complete loss of mTOR activity, leading to a delay in autophagy. PARP-1 depletion favors apoptosis in starved cells, suggesting a pro-survival role of autophagy and PARP-1 activation after nutrient deprivation. In vivo results show that neonates of PARP-1 mutant mice subjected to acute starvation, also display deficient liver autophagy, implying a physiological role for PARP-1 in starvation-induced autophagy. Thus, the PARP signaling pathway is a key regulator of the initial steps of autophagy commitment following starvation.JMRV is recipient of a predoctoral fellowship financed by the program JAE-Pre of CSIC and was also funded by CSIC with a Short-Term Fellowship to stay at the Danish Cancer Society Institute of Cancer Biology in Copenhagen, Denmark. This work was supported by Ministerio de Ciencia e Innovación (SAF2006-01094 and SAF2009-13281-C02-01), Fundación La Caixa (BM06- 219-0) and Junta de Andalucía (P07-CTS-0239) to FJO; RTICC (RD06/0020/0068) to ALR.Peer Reviewe

Interaction between PARP-1 and HIF-2 in the hypoxic response

et al.Hypoxia-inducible factors (HIFs) mediate the transcriptional adaptation of hypoxic cells. The extensive transcriptional programm regulated by HIFs involves the induction of genes controlling angiogenesis, cellular metabolism, cell growth, metastasis, apoptosis, extracellular matrix remodeling and others. HIF is a heterodimer of HIF- and HIF-β subunits. In addition to HIF-1, HIF-2 has evolved as an isoform that contributes differently to the hypoxic adaptation by performing non-redundant functions. Poly (ADP-ribose) polymerase-1 (PARP-1) is a nuclear protein involved in the control of DNA repair and gene transcription by modulating chromatin structure and acting as part of gene-specific enhancer/promoter-binding complexes. Previous results have shown that PARP-1 regulates HIF-1 activity. In this study, we focused on the cross-talk between HIF-2 and PARP-1. By using different approaches to suppress PARP-1, we show that HIF-2 mRNA expression, protein levels and HIF-2-dependent gene expression, such as ANGPTL4 and erythropoietin (EPO), are regulated by PARP-1. This regulation occurs at both the transcriptional and post-trancriptional level. We also show a complex formation between HIF-2 with PARP-1. This complex is sensitive to PARP inhibition and seems to protect against the von Hippel-Lindau-dependent HIF-2 degradation. Finally, we show that parp-1-/-mice display a significant reduction in the circulating hypoxia-induced EPO levels, number of red cells and hemoglobin concentration. Altogether, these results reveal a complex functional interaction between PARP-1 and the HIF system and suggest that PARP-1 is involved in the fine tuning of the HIF-mediated hypoxic response in vivo.This work was supported by Ministerio de Ciencia e Innovación (SAF2006-01094 and SAF2009-13281-C02-01), Fundación La Caixa (BM06-219-0) and Junta de Andalucía (P07-CTS-0239) to FJO; Ministerio de Educación y Ciencia (SAF2007-64597 and SAF-2010-20067) and the BIZKAIA XEDE Program from the Bizkaia County to EB.Peer Reviewe

ROS-induced DNA damage and PARP-1 are required for optimal induction of starvation-induced autophagy

In response to nutrient stress, cells start an autophagy program that can lead to adaptation or death. The mechanisms underlying the signaling from starvation to the initiation of autophagy are not fully understood. In the current study we show that the absence or inactivation of PARP-1 strongly delays starvation-induced autophagy. We have found that DNA damage is an early event of starvation-induced autophagy as measured by γ-H2AX accumulation and comet assay, with PARP-1 knockout cells displaying a reduction in both parameters. During starvation, ROS-induced DNA damage activates PARP-1, leading to ATP depletion (an early event after nutrient deprivation). The absence of PARP-1 blunted AMPK activation and prevented the complete loss of mTOR activity, leading to a delay in autophagy. PARP-1 depletion favors apoptosis in starved cells, suggesting a pro-survival role of autophagy and PARP-1 activation after nutrient deprivation. In vivo results show that neonates of PARP-1 mutant mice subjected to acute starvation, also display deficient liver autophagy, implying a physiological role for PARP-1 in starvation-induced autophagy. Thus, the PARP signaling pathway is a key regulator of the initial steps of autophagy commitment following starvation

ATM form a complex with PARP-1 in DNA that is much more evident after treatment with DNA damaging agents

<p><b>Copyright information:</b></p><p>Taken from "Interaction between ATM and PARP-1 in response to DNA damage and sensitization of ATM deficient cells through PARP inhibition"</p><p>http://www.biomedcentral.com/1471-2199/8/29</p><p>BMC Molecular Biology 2007;8():29-29.</p><p>Published online 25 Apr 2007</p><p>PMCID:PMC1868035.</p><p></p> A: ATM was immunoprecipitated in human melanoma cell lines G361 and HT144 (ATM deficient) as explained under methods and the presence of PARP-1 was tested by immunoblot analysis. Cells were treated with 10 Gy IR or 2 mM MNU for 30 min. In the left panel, PARP-1 was immunoprecipitated from G361 cells treated or not with 2 mM MNU and western blot was performed to reveal ATM. B: Double indirect immunofluorescence in 3T3 fibroblasts (+/+)of PARP-1 (red signal) and ATM (green). Yellow signal correspond with co-localization of both proteins

PARP-1 regulates metastatic melanoma through modulation of vimentin-induced malignant transformation

PARP inhibition can induce anti-neoplastic effects when used as monotherapy or in combination with chemo- or radiotherapy in various tumor settings; however, the basis for the anti-metastasic activities resulting from PARP inhibition remains unknown. PARP inhibitors may also act as modulators of tumor angiogenesis. Proteomic analysis of endothelial cells revealed that vimentin, an intermediary filament involved in angiogenesis and a specific hallmark of EndoMT (endothelial to mesenchymal transition) transformation, was down-regulated following loss of PARP-1 function in endothelial cells. VE-cadherin, an endothelial marker of vascular normalization, was up-regulated in HUVEC treated with PARP inhibitors or following PARP-1 silencing; vimentin over-expression was sufficient to drive to an EndoMT phenotype. In melanoma cells, PARP inhibition reduced pro-metastatic markers, including vasculogenic mimicry. We also demonstrated that vimentin expression was sufficient to induce increased mesenchymal/pro-metastasic phenotypic changes in melanoma cells, including ILK/GSK3-β-dependent E-cadherin down-regulation, Snail1 activation and increased cell motility and migration. In a murine model of metastatic melanoma, PARP inhibition counteracted the ability of melanoma cells to metastasize to the lung. These results suggest that inhibition of PARP interferes with key metastasis-promoting processes, leading to suppression of invasion and colonization of distal organs by aggressive metastatic cells.This work was supported by Ministerio de Ciencia e Innovación SAF2006-01094, SAF2009-13281-C02-01, Fundación La Caixa BM06-219-0 and Junta de Andalucía P07-CTS-0239 and CTS-6602 to FJO, Ministerio de Educación y Ciencia SAF2007-64597; CICYT: SAF2009-13281-C02-02; Junta de Andalucía, P06-CTS-01385 to JMRdA and grants CEIC (P10-CTS5865) and FEDER-ISCIII (PI10/00883) to JCR-M. AGdH has been funded by grants from ‘‘Fundación Científica de la Asociación Española Contra el Cáncer’’, Ministerio de Ciencia y Tecnología SAF2010-16089, and ‘‘Fundación La Marató de TV3’’. JCR-M has been funded by Grants CEIC (P1=-CTS5865) and FEDER-ISCIII (PI10/00883). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscriptBaulida and García de Herreros' labs is supported by the Instituto de Salud Carlos III (PI12/ 00257 and D012/0036/0005, part of the Plan Nacional I+D+ I and cofounded by the ISCIII-Subdirección General de Evaluación and Fondo Europeo de Desarrollo Regional-FEDER), the Fundación Científica Asociación Española Contra el Cáncer (Ayudas a grupos estables de investigación, 2010-1) and the Ministerio de Economía y Competitividad (SAF2013-4889-C2-1R)