PARP1
Poly [ADP-ribose] polymerase 1 (EC 2.4.2.30) (PARP-1) (ADP-ribosyltransferase diphtheria toxin-like 1) (ARTD1) (DNA ADP-ribosyltransferase PARP1) (EC 2.4.2.-) (NAD( ) ADP-ribosyltransferase 1) (ADPRT 1) (Poly[ADP-ribose] synthase 1) (Protein poly-ADP-ribosyltransferase PARP1) (EC 2.4.2.-) [ADPRT] [PPOL]
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Leukocyte telomere length (LTL) is a heritable biomarker of genomic aging. In this study, we perform a genome-wide meta-analysis of LTL by pooling densely genotyped and imputed association results across large-scale European-descent studies including up to 78,592 individuals. We identify 49 genomic regions at a false dicovery rate (FDR) < 0.05 threshold and prioritize genes at 31, with five highlighting nucleotide metabolism as an important regulator of LTL. We report six genome-wide significant loci in or near SENP7, MOB1B, CARMIL1, PRRC2A, TERF2, and RFWD3, and our results support recently identified PARP1, POT1, ATM, and MPHOSPH6 loci. Phenome-wide analyses in >350,000 UK Biobank participants suggest that genetically shorter telomere length increases the risk of hypothyroidism and decreases the risk of thyroid cancer, lymphoma, and a range of proliferative conditions. Our results replicate previously reported associations with increased risk of coronary artery disease and lower risk for multiple cancer types. Our findings substantially expand current knowledge on genes that regulate LTL and their impact on human health and disease.
MeSH Terms
- Genome-Wide Association Study
- Humans
- Leukocytes
- Nucleotides
- Telomere
Keywords
- Mendelian randomisation
- age-related disease
- biological aging
- telomere length
Nicotinamide adenine dinucleotide (NAD) is an important coenzyme that regulates various metabolic pathways, including glycolysis, β-oxidation, and oxidative phosphorylation. Additionally, NAD serves as a substrate for poly(ADP-ribose) polymerase (PARP), sirtuin, and NAD glycohydrolase, and it regulates DNA repair, gene expression, energy metabolism, and stress responses. Many studies have demonstrated that NAD metabolism is deeply involved in aging and aging-related diseases. Previously, we demonstrated that nicotinamide guanine dinucleotide (NGD) and nicotinamide hypoxanthine dinucleotide (NHD), which are analogs of NAD, are significantly increased in Nmnat3-overexpressing mice. However, there is insufficient knowledge about NGD and NHD in vivo. In the present study, we aimed to investigate the metabolism and biochemical properties of these NAD analogs. We demonstrated that endogenous NGD and NHD were found in various murine tissues, and their synthesis and degradation partially rely on Nmnat3 and CD38. We have also shown that NGD and NHD serve as coenzymes for alcohol dehydrogenase (ADH) in vitro, although their affinity is much lower than that of NAD. On the other hand, NGD and NHD cannot be used as substrates for SIRT1, SIRT3, and PARP1. These results reveal the basic metabolism of NGD and NHD and also highlight their biological function as coenzymes.
MeSH Terms
- Aging
- Animals
- Guanine Nucleotides
- Guanosine Triphosphate
- Inosine Triphosphate
- Mice
- NAD
- Poly(ADP-ribose) Polymerases
- Sirtuins
T cells play a key role in controlling viral infections; however, the underlying mechanisms regulating their functions during human viral infections remain incompletely understood. Here, we used CD4 T cells derived from individuals with chronic viral infections or healthy T cells treated with camptothecin (CPT) - a topoisomerase I (Top 1) inhibitor - as a model to investigate the role of DNA topology in reprogramming telomeric DNA damage responses (DDR) and remodeling T cell functions. We demonstrated that Top 1 protein expression and enzyme activity were significantly inhibited, while the Top 1 cleavage complex (TOP1cc) was trapped in genomic DNA, in T cells derived from individuals with chronic viral (HCV, HBV, or HIV) infections. Top 1 inhibition by CPT treatment of healthy CD4 T cells caused topological DNA damage, telomere attrition, and T cell apoptosis or dysfunction via inducing Top1cc accumulation, PARP1 cleavage, and failure in DNA repair, thus recapitulating T cell dysregulation in the setting of chronic viral infections. Moreover, T cells from virally infected subjects with inhibited Top 1 activity were more vulnerable to CPT-induced topological DNA damage and cell apoptosis, indicating an important role for Top 1 in securing DNA integrity and cell survival. These findings provide novel insights into the molecular mechanisms for immunomodulation by chronic viral infections via disrupting DNA topology to induce telomeric DNA damage, T cell senescence, apoptosis and dysfunction. As such, restoring the impaired DNA topologic machinery may offer a new strategy for maintaining T cell function against human viral diseases.
Keywords
- HBV
- HCV
- HIV
- T cell senescence
- Telomere attrition
- Topological DNA damage
Nicotinamide adenine dinucleotide (NAD ) is a critical molecule involved in various biological functions. Poly (ADP-ribose) polymerase 1 (PARP1) and sirtuin 1 (SIRT1) affect cellular NAD levels and play essential roles in regulating metabolism. However, there has been little research on the effects of PARP1 and SIRT1 crosstalk during senescence. We isolated endothelial progenitor cells (EPCs) from human umbilical cord blood and treated them with a PARP1 inhibitor (PJ34). Using a stress-induced premature aging model built by H O , transfection with adenoviral vectors, and Western blot analysis, we observed that PJ34 treatment preserved intracellular NAD levels, increased SIRT1 activity, decreased p53 acetylation, and improved the function of stress-induced premature aging EPCs. Our results suggest that PJ34 improves the function of aging-induced EPCs and may contribute to cellular therapies for atherosclerosis.
MeSH Terms
- Cell Movement
- Cellular Senescence
- Cyclin-Dependent Kinase Inhibitor p21
- Endothelial Progenitor Cells
- Fetal Blood
- Gene Expression Regulation
- Humans
- Hydrogen Peroxide
- NAD
- Oxidative Stress
- Phenanthrenes
- Poly (ADP-Ribose) Polymerase-1
- Primary Cell Culture
- Signal Transduction
- Sirtuin 1
- Tumor Suppressor Protein p53
Keywords
- Endothelial progenitor cells
- Nicotinamide adenine dinucleotide
- Poly (ADP-ribose) polymerase 1
- Senescence
- Sirtuin 1
Mitochondrial dysfunction, inflammation and senescence-like features are observed in adipose depots in aging and obesity. Herein, we evaluated how maternal high calorie diet (HCD) may impact on subcutaneous adipose tissue (sAT) of the newborn mice. Adult C57BL/6J mice were randomly divided in three groups: normal calorie diet (NCD), HCD and HCD supplemented with niacin 8 weeks before mating. Mothers and pups were then sacrificed and metabolic and molecular analyses were carried out on sAT. HCD induced mitochondria dysfunction in mothers without inflammation and senescence, whereas in pups we also revealed the occurrence of senescent phenotype. The mitochondrial dysfunction-associated senescence in pups was accompanied by a drop in NAD /NADH ratio and alteration in the NAD -dependent enzymes PARP1 and SIRT1. Importantly, maternal dietary supplementation with niacin during gestation and lactation restrained NAD /NADH decrease imposed by HCD limiting inflammatory cytokine production and senescence phenotype in newborn sAT. Given the fundamental role of sAT in buffering nutrient overload and avoiding pathogenic ectopic fat accumulation, we suggest that NAD boosting strategies during maternal HCD could be helpful in limiting sAT dysfunction in newborn.
Keywords
- Gerotarget
- NAD
- adipocytes
- aging
- inflammation
- senescence
Loss of function of dyskerin (DKC1), NOP10 and TIN2 are responsible for different inheritance patterns of Dyskeratosis congenita (DC; ORPHA1775). They are key components of telomerase (DKC1 and NOP10) and shelterin (TIN2), and play an important role in telomere homeostasis. They participate in several fundamental cellular processes by contributing to Dyskeratosis congenita through mechanisms that are not fully understood. Presence of oxidative stress was postulated to result from telomerase ablation. However, the resulting disturbed redox status can promote telomere attrition by generating a vicious circle, which promotes cellular senescence. This fact prompted us to study if acute loss of DKC1, NOP10 and TINF2 can promote redox disequilibrium as an early event when telomere shortening has not yet taken place. We generated siRNA-mediated (DKC1, NOP10 and TINF2) cell lines by RNA interference, which was confirmed by mRNA and protein expression analyses. No telomere shortening occurred in any silenced cell line. Depletion of H/ACA ribonucleoproteins DKC1 and NOP10 diminished telomerase activity via TERC down-regulation, and produced alterations in pseudouridylation and ribosomal biogenesis. An increase in the GSSG/GSH ratio, carbonylated proteins and oxidized peroxiredoxin-6 was observed, in addition to MnSOD and TRX1 overexpression in the siRNA DC cells. Likewise, high PARylation levels and high PARP1 protein expression were detected. In contrast, the silenced TINF2 cells did not alter any evaluated oxidative stress marker. Altogether these findings lead us to conclude that loss of DKC1 and NOP10 functions induces oxidative stress in a telomere shortening independent manner.
MeSH Terms
- Cell Cycle Proteins
- Cellular Senescence
- DNA Damage
- Dyskeratosis Congenita
- HeLa Cells
- Humans
- Nuclear Proteins
- Oxidative Stress
- RNA Interference
- Ribonucleoproteins, Small Nucleolar
- Telomerase
- Telomere Shortening
- Telomere-Binding Proteins
Keywords
- Aging
- Antioxidant
- DNA damage
- Oxidative stress
- Telomeropathies
A deeper understanding of the conserved molecular mechanisms in different taxa have been made possible only because of the evolutionary conservation of crucial signaling pathways. In the present study, we explored the molecular evolutionary pattern of selection signatures in 51 species for 10 genes which are important components of NAD /Sirtuin pathway and have already been directly linked to lifespan extension in worms and mice. Selection pressure analysis using PAML program revealed that MRPS5 and PPARGC1A were under significant constraints because of their functional significance. FOXO3a also displayed strong purifying selection. All three sirtuins, which were SIRT1, SIRT2 and SIRT6, displayed a great degree of conservation between taxa, which is consistent with the previous report. A significant evolutionary constraint is seen on the anti-oxidant gene, SOD3. As expected, TP53 gene was under significant selection pressure in mammals, owing to its major role in tumor progression. Poly-ADP-ribose polymerase (PARP) genes displayed the most sites under positive selection. Further 3D structural analysis of PARP1 and PARP2 protein revealed that some of these positively selected sites caused a change in the electrostatic potential of the protein structure, which may allow a change in its interaction with other proteins and molecules ultimately leading to difference in the function. Although the functional significance of the positively selected sites could not be established in the variants databases, yet it will be interesting to see if these sites actually affect the function of PARP1 and PARP2.
MeSH Terms
- Aging
- Animals
- Evolution, Molecular
- Helminths
- Humans
- Mice
- NAD
- Poly (ADP-Ribose) Polymerase-1
- Poly(ADP-ribose) Polymerases
- Selection, Genetic
- Signal Transduction
- Sirtuins
NAD levels decline with age in diverse animals from Caenorhabditis elegans to mice. Raising NAD levels by dietary supplementation with NAD precursors, nicotinamide riboside (NR) or nicotinamide mononucleotide (NMN), improves mitochondrial function and muscle and neural and melanocyte stem cell function in mice, as well as increases murine life span. Decreased NAD levels with age reduce SIRT1 function and reduce the mitochondrial unfolded protein response, which can be overcome by NR supplementation. Decreased NAD levels cause NAD -binding protein DBC1 to form a complex with PARP1, inhibiting poly(adenosine diphosphate-ribose) polymerase (PARP) catalytic activity. Old mice have increased amounts of DBC1-PARP1 complexes, lower PARP activity, increased DNA damage, and reduced nonhomologous end joining and homologous recombination repair. DBC1-PARP1 complexes in old mice can be broken by increasing NAD levels through treatment with NMN, reducing DNA damage and restoring PARP activity to youthful levels. The mechanism of declining NAD levels and its fundamental importance to aging are yet to be elucidated. There is a correlation of PARP activity with mammalian life span that suggests that NAD /SIRT1/PARP1 may be more significant than the modest effects on life span observed for NR supplementation in old mice. The NAD /PARP1/SIRT1 axis may link NAD levels and DNA damage with the apparent epigenomic DNA methylation clocks that have been described.
MeSH Terms
- Aging
- Animals
- DNA Damage
- Humans
- Mice
- NAD
- Poly(ADP-ribose) Polymerases
- Signal Transduction
- Sirtuin 1
Keywords
- DBC1/CCAR2
- NAD
- PARP1
- SIRT1
- epigenome
DNA repair is essential for life, yet its efficiency declines with age for reasons that are unclear. Numerous proteins possess Nudix homology domains (NHDs) that have no known function. We show that NHDs are NAD (oxidized form of nicotinamide adenine dinucleotide) binding domains that regulate protein-protein interactions. The binding of NAD to the NHD domain of DBC1 (deleted in breast cancer 1) prevents it from inhibiting PARP1 [poly(adenosine diphosphate-ribose) polymerase], a critical DNA repair protein. As mice age and NAD concentrations decline, DBC1 is increasingly bound to PARP1, causing DNA damage to accumulate, a process rapidly reversed by restoring the abundance of NAD Thus, NAD directly regulates protein-protein interactions, the modulation of which may protect against cancer, radiation, and aging.
MeSH Terms
- Adaptor Proteins, Signal Transducing
- Aging
- Animals
- Conserved Sequence
- DNA Damage
- DNA Repair
- Fibroblasts
- HEK293 Cells
- Humans
- Mice
- Models, Molecular
- NAD
- Neoplasms
- Paraquat
- Poly (ADP-Ribose) Polymerase-1
- Protein Interaction Domains and Motifs
- RNA, Small Interfering
- Radiation Tolerance
- Sequence Homology, Nucleic Acid
Cellular senescence is an important tumor-suppressive mechanism. However, acquisition of a senescence-associated secretory phenotype (SASP) in senescent cells has deleterious effects on the tissue microenvironment and, paradoxically, promotes tumor progression. In a drug screen, we identified melatonin as a novel SASP suppressor in human cells. Strikingly, melatonin blunts global SASP gene expression upon oncogene-induced senescence (OIS). Moreover, poly(ADP-ribose) polymerase-1 (PARP-1), a sensor of DNA damage, was identified as a new melatonin-dependent regulator of SASP gene induction upon OIS. Here, we report two different but potentially coherent epigenetic strategies for melatonin regulation of SASP. The interaction between the telomeric repeat-containing RNA (TERRA) and PARP-1 stimulates the SASP, which was attenuated by 67.9% (illustrated by the case of IL8) by treatment with melatonin. Through binding to macroH2A1.1, PARP-1 recruits CREB-binding protein (CBP) to mediate acetylation of H2BK120, which positively regulates the expression of target SASP genes, and this process is interrupted by melatonin. Consequently, the findings provide novel insight into melatonin's epigenetic role via modulating PARP-1 in suppression of SASP gene expression in OIS-induced senescent cells. Our studies identify melatonin as a novel anti-SASP molecule, define PARP-1 as a new target by which melatonin regulates SASP, and establish a new epigenetic paradigm for a pharmacological mechanism by which melatonin interrupts PARP-1 interaction with the telomeric long noncoding RNA(lncRNA) or chromatin.
MeSH Terms
- Cell Line
- Cells, Cultured
- Cellular Senescence
- Fibroblasts
- Humans
- Lung
- Melatonin
- Poly (ADP-Ribose) Polymerase-1
Keywords
SASP
TERRA
- PARP1
- cellular senescence
- histone acetylation
- melatonin
As senescence develops, cells sequentially acquire diverse senescent phenotypes along with simultaneous multistage gene reprogramming. It remains unclear what acts as the key regulator of the collective changes in gene expression at initiation of senescent reprogramming. Here we analyzed time series gene expression profiles obtained in two different senescence models in human diploid fibroblasts: replicative senescence and H O -induced senescence. Our results demonstrate that suppression of DNA methyltransferase 1 (DNMT1)-mediated DNA methylation activity was an initial event prior to the display of senescent phenotypes. We identified seven DNMT1-interacting proteins, ubiquitin-like with PHD and ring finger domains 1 (UHRF1), EZH2, CHEK1, SUV39H1, CBX5, PARP1, and HELLS (also known as LSH (lymphoid-specific helicase) 1), as being commonly down-regulated at the same time point as DNMT1 in both senescence models. Knockdown experiments revealed that, among the DNMT1-interacting proteins, only UHRF1 knockdown suppressed DNMT1 transcription. However, UHRF1 overexpression alone did not induce DNMT1 expression, indicating that UHRF1 was essential but not sufficient for DNMT1 transcription. Although UHRF1 knockdown effectively induced senescence, this was significantly attenuated by DNMT1 overexpression, clearly implicating the UHRF1/DNMT1 axis in senescence. Bioinformatics analysis further identified WNT5A as a downstream effector of UHRF1/DNMT1-mediated senescence. Senescence-associated hypomethylation was found at base pairs -1569 to -1363 from the transcription start site of the WNT5A gene in senescent human diploid fibroblasts. As expected, WNT5A overexpression induced senescent phenotypes. Overall, our results indicate that decreased UHRF1 expression is a key initial event in the suppression of DNMT1-mediated DNA methylation and in the consequent induction of senescence via increasing WNT5A expression.
MeSH Terms
- CCAAT-Enhancer-Binding Proteins
- Cellular Senescence
- DNA (Cytosine-5-)-Methyltransferase 1
- DNA (Cytosine-5-)-Methyltransferases
- DNA Methylation
- Fibroblasts
- Gene Expression Profiling
- Gene Expression Regulation
- HEK293 Cells
- Histones
- Humans
- Hydrogen Peroxide
- Male
- Oligonucleotide Array Sequence Analysis
- Phenotype
- Promoter Regions, Genetic
- Protein Binding
- Protein Domains
- RNA, Small Interfering
- Ubiquitin-Protein Ligases
- Wnt-5a Protein
- beta-Galactosidase
Keywords
- DNA methylation
- cellular senescence
- gene expression
- gene regulation
- microarray
In [i]C. elegans[/i], miRNAs are genetic biomarkers of aging. Similarly, multiple miRNAs are differentially expressed between younger and older persons, suggesting that miRNA-regulated biological mechanisms affecting aging are evolutionarily conserved. Previous human studies have not considered participants' lifespans, a key factor in identifying biomarkers of aging. Using PCR arrays, we measured miRNA levels from serum samples obtained longitudinally at ages 50, 55, and 60 from 16 non-Hispanic males who had documented lifespans from 58 to 92. Numerous miRNAs showed significant changes in expression levels. At age 50, 24 miRNAs were significantly upregulated, and 73 were significantly downregulated in the long-lived subgroup (76-92 years) as compared with the short-lived subgroup (58-75 years). In long-lived participants, the most upregulated was miR-373-5p, while the most downregulated was miR-15b-5p. Longitudinally, significant Pearson correlations were observed between lifespan and expression of nine miRNAs (p value<0.05). Six of these nine miRNAs (miR-211-5p, 374a-5p, 340-3p, 376c-3p, 5095, 1225-3p) were also significantly up- or downregulated when comparing long-lived and short-lived participants. Twenty-four validated targets of these miRNAs encoded aging-associated proteins, including PARP1, IGF1R, and IGF2R. We propose that the expression profiles of the six miRNAs (miR-211-5p, 374a-5p, 340-3p, 376c-3p, 5095, and 1225-3p) may be useful biomarkers of aging.
MeSH Terms
- Aged
- Aged, 80 and over
- Aging
- Animals
- Biomarkers
- Caenorhabditis elegans
- Down-Regulation
- Gene Expression Profiling
- Humans
- Longevity
- Longitudinal Studies
- Male
- MicroRNAs
- Middle Aged
- Pilot Projects
- Polymerase Chain Reaction
- Up-Regulation
Keywords
- aging
- biomarker
- long-lived
- longitudinal study
- miRNA
- short-lived
Cockayne syndrome is a neurodegenerative accelerated aging disorder caused by mutations in the CSA or CSB genes. Although the pathogenesis of Cockayne syndrome has remained elusive, recent work implicates mitochondrial dysfunction in the disease progression. Here, we present evidence that loss of CSA or CSB in a neuroblastoma cell line converges on mitochondrial dysfunction caused by defects in ribosomal DNA transcription and activation of the DNA damage sensor poly-ADP ribose polymerase 1 (PARP1). Indeed, inhibition of ribosomal DNA transcription leads to mitochondrial dysfunction in a number of cell lines. Furthermore, machine-learning algorithms predict that diseases with defects in ribosomal DNA (rDNA) transcription have mitochondrial dysfunction, and, accordingly, this is found when factors involved in rDNA transcription are knocked down. Mechanistically, loss of CSA or CSB leads to polymerase stalling at non-B DNA in a neuroblastoma cell line, in particular at G-quadruplex structures, and recombinant CSB can melt G-quadruplex structures. Indeed, stabilization of G-quadruplex structures activates PARP1 and leads to accelerated aging in Caenorhabditis elegans In conclusion, this work supports a role for impaired ribosomal DNA transcription in Cockayne syndrome and suggests that transcription-coupled resolution of secondary structures may be a mechanism to repress spurious activation of a DNA damage response.
MeSH Terms
- Cell Line, Tumor
- Cockayne Syndrome
- DNA Damage
- DNA Helicases
- DNA Repair
- DNA Repair Enzymes
- DNA, Neoplasm
- DNA, Ribosomal
- G-Quadruplexes
- Gene Knockdown Techniques
- Humans
- Neuroblastoma
- Poly (ADP-Ribose) Polymerase-1
- Poly-ADP-Ribose Binding Proteins
- Transcription Factors
- Transcription, Genetic
Keywords
- CSA
- CSB
- Cockayne syndrome
- aging
- nucleolus
- polymerase I transcription
Recently, we demonstrated the association of sperm-associated antigen 9 (SPAG9) expression with breast cancer. Among breast cancer, 15 % of the cancers are diagnosed as triple-negative breast cancers (TNBC) based on hormone receptor status and represent an important clinical challenge because of lack of effective available targeted therapy. Therefore, in the present investigation, plasmid-based small hairpin (small hairpin RNA (shRNA)) approach was used to ablate SPAG9 in aggressive breast cancer cell line model (MDA-MB-231) in order to understand the role of SPAG9 at molecular level in apoptosis, cell cycle, and epithelial-to-mesenchymal transition (EMT) signaling. Our data in MDA-MB-231 cells showed that ablation of SPAG9 resulted in membrane blebbing, increased mitochondrial membrane potential, DNA fragmentation, phosphatidyl serine surface expression, and caspase activation. SPAG9 depletion also resulted in cell cycle arrest in G0-G1 phase and induced cellular senescence. In addition, in in vitro and in vivo xenograft studies, ablation of SPAG9 resulted in upregulation of p21 along with pro-apoptotic molecules such as BAK, BAX, BIM, BID, NOXA, AIF, Cyto-C, PARP1, APAF1, Caspase 3, and Caspase 9 and epithelial marker, E-cadherin. Also, SPAG9-depleted cells showed downregulation of cyclin B1, cyclin D1, cyclin E, CDK1, CDK4, CDK6, BCL2, Bcl-xL, XIAP, cIAP2, MCL1, GRP78, SLUG, SNAIL, TWIST, vimentin, N-cadherin, MMP2, MMP3, MMP9, SMA, and β-catenin. Collectively, our data suggests that SPAG9 promotes tumor growth by inhibiting apoptosis, altering cell cycle, and enhancing EMT signaling in in vitro cells and in vivo mouse model. Hence, SPAG9 may be a potential novel target for therapeutic use in TNBC treatment.
MeSH Terms
- Adaptor Proteins, Signal Transducing
- Animals
- Apoptosis
- Blotting, Western
- Cell Proliferation
- Fluorescent Antibody Technique, Indirect
- Humans
- Immunoenzyme Techniques
- Membrane Potential, Mitochondrial
- Mice
- RNA, Small Interfering
- Triple Negative Breast Neoplasms
- Tumor Cells, Cultured
Keywords
- Apoptosis
- Cell growth
- Cellular motility
- SPAG9
- Senescence
- Triple-negative breast cancer
- Tumor growth
Chemotherapy is one of the major treatment modalities for cancer. Metal-based compounds such as derivatives of cisplatin are in the front line of therapy against a subset of cancers, but their use is restricted by severe side-effects and the induction of resistance in treated tumors. Subsequent research focused on development of cytotoxic metal-complexes without cross-resistance to cisplatin and reduced side-effects. This led to the discovery of first-generation titanium(IV)salan complexes, which reached clinical trials but lacked efficacy. New-generation titanium (IV)salan-complexes show promising anti-tumor activity in mice, but their molecular mechanism of cytotoxicity is completely unknown. Four different human cell lines were analyzed in their responses to a toxic (Tc52) and a structurally highly related but non-toxic (Tc53) titanium(IV)salan complex. Viability assays were used to reveal a suitable treatment range, flow-cytometry analysis was performed to monitor the impact of dosage and treatment time on cell-cycle distribution and cell death. Potential DNA strand break induction and crosslinking was investigated by immunostaining of damage markers as well as automated fluorometric analysis of DNA unwinding. Changes in nuclear morphology were analyzed by DAPI staining. Acidic beta-galactosidase activity together with morphological changes was monitored to detect cellular senescence. Western blotting was used to analyze induction of pro-apoptotic markers such as activated caspase7 and cleavage of PARP1, and general stress kinase p38. Here we show that the titanium(IV)salan Tc52 is effective in inducing cell death in the lower micromolar range. Surprisingly, Tc52 does not target DNA contrary to expectations deduced from the reported activity of other titanium complexes. Instead, Tc52 application interferes with progression from G2-phase into mitosis and induces apoptotic cell death in tested tumor cells. Contrarily, human fibroblasts undergo senescence in a time and dose-dependent manner. As deduced from fluorescence studies, the potential cellular target seems to be the cytoskeleton. In summary, we could demonstrate in four different human cell lines that tumor cells were specifically killed without induction of major cytotoxicity in non-tumorigenic cells. Absence of DNA damaging activity and the cell-cycle block in G2 instead of mitosis makes Tc52 an attractive compound for further investigations in cancer treatment.
MeSH Terms
- Antineoplastic Agents
- Apoptosis
- Blotting, Western
- Caspase 7
- Cell Division
- Cell Line, Tumor
- Cell Nucleus
- Cell Survival
- Cellular Senescence
- Coordination Complexes
- Cytoskeleton
- DNA Damage
- Flow Cytometry
- G2 Phase
- HEK293 Cells
- HeLa Cells
- Humans
- Poly (ADP-Ribose) Polymerase-1
- Radioisotopes
- Titanium
- p38 Mitogen-Activated Protein Kinases
Keywords
- Apoptosis
- Cell-cycle
- Senescence
- Titanium(IV)salan complex
- Tumorigenicity
Hutchinson Gilford progeria syndrome is a fatal disorder characterized by accelerated aging, bone resorption and atherosclerosis, caused by a LMNA mutation which produces progerin, a mutant lamin A precursor. Progeria cells display progerin and prelamin A nuclear accumulation, altered histone methylation pattern, heterochromatin loss, increased DNA damage and cell cycle alterations. Since the LMNA promoter contains a retinoic acid responsive element, we investigated if all-trans retinoic acid administration could lower progerin levels in cultured fibroblasts. We also evaluated the effect of associating rapamycin, which induces autophagic degradation of progerin and prelamin A. We demonstrate that all-trans retinoic acid acts synergistically with low-dosage rapamycin reducing progerin and prelamin A, via transcriptional downregulation associated with protein degradation, and increasing the lamin A to progerin ratio. These effects rescue cell dynamics and cellular proliferation through recovery of DNA damage response factor PARP1 and chromatin-associated nuclear envelope proteins LAP2α and BAF. The combined all-trans retinoic acid-rapamycin treatment is dramatically efficient, highly reproducible, represents a promising new approach in Hutchinson-Gilford Progeria therapy and deserves investigation in ageing-associated disorders.
MeSH Terms
- Antineoplastic Agents
- Blotting, Western
- Cell Cycle
- Cell Proliferation
- Cells, Cultured
- DNA-Binding Proteins
- Drug Synergism
- Fibroblasts
- Gene Expression
- Histones
- Humans
- Lamin Type A
- Lysine
- Membrane Proteins
- Methylation
- Microscopy, Electron, Transmission
- Microscopy, Fluorescence
- Nuclear Proteins
- Phenotype
- Poly (ADP-Ribose) Polymerase-1
- Poly(ADP-ribose) Polymerases
- Progeria
- Reverse Transcriptase Polymerase Chain Reaction
- Sirolimus
- Tretinoin
Keywords
- DNA damage and repair
- Hutchinson Gilford progeria syndrome
- all-trans retinoic acid
- premature aging
- rapamycin
In principle, a decline in base excision repair (BER) efficiency with age should lead to genomic instability and ultimately contribute to the onset of the aging phenotype. Although multiple studies have indicated a negative link between aging and BER, the change of BER efficiency with age in humans has not been systematically analyzed. Here, with foreskin fibroblasts isolated from 19 donors between 20 and 64 y of age, we report a significant decline of BER efficiency with age using a newly developed GFP reactivation assay. We further observed a very strong negative correlation between age and the expression levels of SIRT6, a factor which is known to maintain genomic integrity by improving DNA double strand break (DSB) repair. Our mechanistic study suggests that, similar to the regulatory role that SIRT6 plays in DNA DSB repair, SIRT6 regulates BER in a PARP1-depdendent manner. Moreover, overexpression of SIRT6 rescues the decline of BER in aged fibroblasts. In summary, our results uncovered the regulatory mechanisms of BER by SIRT6, suggesting that SIRT6 reactivation in aging tissues may help delay the process of aging through improving BER.
MeSH Terms
- Adult
- Aging
- Cells, Cultured
- DNA Breaks, Double-Stranded
- DNA Repair
- Fibroblasts
- Foreskin
- Genomic Instability
- Humans
- Light
- Male
- Middle Aged
- Phenanthrenes
- Poly(ADP-ribose) Polymerase Inhibitors
- Poly(ADP-ribose) Polymerases
- RNA Interference
- RNA, Small Interfering
- Sirtuins
Keywords
- PARP1
- SIRT6
- SIRTUIN
- aging
- base excision repair
- mono-ADP-ribosylation
Genome instability represents a primary hallmark of aging and cancer. RecQL helicases (i.e., RECQL1, WRN, BLM, RECQL4, RECQL5) as well as poly(ADP-ribose) polymerases (PARPs, in particular PARP1) represent two central quality control systems to preserve genome integrity in mammalian cells. Consistently, both enzymatic families have been linked to mechanisms of aging and carcinogenesis in mice and humans. This is in accordance with clinical and epidemiological findings demonstrating that defects in three RecQL helicases, i.e., WRN, BLM, RECQL4, are related to human progeroid and cancer predisposition syndromes, i.e., Werner, Bloom, and Rothmund Thomson syndrome, respectively. Moreover, PARP1 hypomorphy is associated with a higher risk for certain types of cancer. On a molecular level, RecQL helicases and PARP1 are involved in the control of DNA repair, telomere maintenance, and replicative stress. Notably, over the last decade, it became apparent that all five RecQL helicases physically or functionally interact with PARP1 and/or its enzymatic product poly(ADP-ribose) (PAR). Furthermore, a profound body of evidence revealed that the cooperative function of RECQLs and PARP1 represents an important factor for maintaining genome integrity. In this review, we summarize the status quo of this molecular cooperation and discuss open questions that provide a basis for future studies to dissect the cooperative functions of RecQL helicases and PARP1 in aging and carcinogenesis.
MeSH Terms
- Aging
- Animals
- DNA
- Genome, Human
- Genomic Instability
- Humans
- Mice
- Poly (ADP-Ribose) Polymerase-1
- Poly(ADP-ribose) Polymerases
- RecQ Helicases
Keywords
- Aging
- BLM
- Cancer
- Poly(ADP-ribose) polymerase
- RECQL
- WRN
Hutchinson-Gilford progeria syndrome (HGPS) is a severe human premature aging disorder caused by a lamin A mutant named progerin. Death occurs at a mean age of 13 y from cardiovascular problems. Previous studies revealed loss of vascular smooth muscle cells (SMCs) in the media of large arteries in a patient with HGPS and two mouse models, suggesting a causal connection between the SMC loss and cardiovascular malfunction. However, the mechanisms of how progerin leads to massive SMC loss are unknown. In this study, using SMCs differentiated from HGPS induced pluripotent stem cells, we show that HGPS SMCs exhibit a profound proliferative defect, which is primarily caused by caspase-independent cell death. Importantly, progerin accumulation stimulates a powerful suppression of PARP1 and consequently triggers an activation of the error-prone nonhomologous end joining response. As a result, most HGPS SMCs exhibit prolonged mitosis and die of mitotic catastrophe. This study demonstrates a critical role of PARP1 in mediating SMC loss in patients with HGPS and elucidates a molecular pathway underlying the progressive SMC loss in progeria.
MeSH Terms
- Aging
- Cell Death
- Cell Differentiation
- Cell Survival
- Down-Regulation
- Fibroblasts
- G2 Phase
- Humans
- Myocytes, Smooth Muscle
- Pluripotent Stem Cells
- Poly (ADP-Ribose) Polymerase-1
- Poly(ADP-ribose) Polymerases
- Primary Cell Culture
- Progeria
- S Phase
- Skin
Sirtuin 1 (SIRT1) regulates inflammation, aging (life span and health span), calorie restriction/energetics, mitochondrial biogenesis, stress resistance, cellular senescence, endothelial functions, apoptosis/autophagy, and circadian rhythms through deacetylation of transcription factors and histones. SIRT1 level and activity are decreased in chronic inflammatory conditions and aging, in which oxidative stress occurs. SIRT1 is regulated by a NAD( )-dependent DNA repair enzyme, poly(ADP-ribose) polymerase-1 (PARP1), and subsequent NAD( ) depletion by oxidative stress may have consequent effects on inflammatory and stress responses as well as cellular senescence. SIRT1 has been shown to undergo covalent oxidative modifications by cigarette smoke-derived oxidants/aldehydes, leading to posttranslational modifications, inactivation, and protein degradation. Furthermore, oxidant/carbonyl stress-mediated reduction of SIRT1 leads to the loss of its control on acetylation of target proteins including p53, RelA/p65, and FOXO3, thereby enhancing the inflammatory, prosenescent, and apoptotic responses, as well as endothelial dysfunction. In this review, the mechanisms of cigarette smoke/oxidant-mediated redox posttranslational modifications of SIRT1 and its roles in PARP1 and NF-κB activation, and FOXO3 and eNOS regulation, as well as chromatin remodeling/histone modifications during inflammaging, are discussed. Furthermore, we have also discussed various novel ways to activate SIRT1 either directly or indirectly, which may have therapeutic potential in attenuating inflammation and premature senescence involved in chronic lung diseases.
MeSH Terms
- Active Transport, Cell Nucleus
- Animals
- Cellular Senescence
- Forkhead Box Protein O3
- Forkhead Transcription Factors
- Histones
- Humans
- Inflammation
- NF-kappa B
- Nitric Oxide Synthase Type III
- Oxidation-Reduction
- Oxidative Stress
- Poly (ADP-Ribose) Polymerase-1
- Poly(ADP-ribose) Polymerases
- Signal Transduction
- Sirtuin 1
- Transcription Factor RelA
Keywords
- COPD
- FOXO3
- Free radicals
- GSH
- Inflammation
- NF-κB
- Oxidants
- Redox signaling
- SIRT1
- Senescence
- Tobacco smoke