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Purpose: Alpha-blockers have been proven as an effective method for increasing the stone expulsion rate of distal ureteral stones. Limited studies have focused on doxazosin; its efficacy remained unclear. We performed this meta-analysis to investigate the efficacy and safety of doxazosin for patients diagnosed with distal ureteral stones less than 10mm.Materials and Methods: We systematically searched Ovid MEDLINE®, Cochrane Library, EMBASE, and PubMed for articles comparing doxazosin and conventional care or tamsulosin for distal ureteral stones through October 2019. The outcome measures were stone expulsive rate (SER), stone expulsive time (SET), pain episodes, analgesics consumption, and adverse events.Results: We included 12 studies involving 836 participants with distal ureteral stones less than 10mm in our review. The present meta-analysis showed doxazosin could significantly increase SER [RR=1.64,95%CI (1.32, 2.04), P  < 0.00001], shorten SET [WMD=-3.97,95% CI (-5.68, -2.27), P  < 0.00001] compared with conventional care. In the subgroup analyses, doxazosin showed no benefit in the children subgroup (<16 years old) [RR=1.63,95% CI (0.73,3.64), P  =0.23]. No statistically significant difference was observed regarding the effectiveness of doxazosin and tamsulosin in SER, SET, and safety. 9 in 286 participants reported doxazosin-related adverse events; most were mild to moderate.Conclusion: This meta-analysis may suggest that doxazosin is a safe and effective MET for distal ureteral stones less than 10mm. It is not demonstrated to have any significant difference with tamsulosin in SER, SET, and safety. However, it showed no benefits for patients<16 years old. [ABSTRACT FROM AUTHOR]

To study the characteristics of tuber starchgranules, firstly, three kinds of common tuber starches were separated and characterized by gravitational field-flow fractionation (GrFFF) instrument, and the ratios of large and small starch granules A/B were calculated. Then, the liquid of peak A and peak B was collected and observed by scanning electron microscope (SEM) to verify the separation result of GrFFF. Secondly, the particle size distribution and crystal characteristics were characterized by particle size analyser, X-ray diffraction (X-ray) and fourier transform infrared spectroscopy (FT-IR). Three kinds of tuber starches all had the bimodal distribution in GrFFF. Peak A contained large particles, and peak B contained small particles. For yam starch, the ratio of starch granules A/B was much higher than 1; for sweet potato starch, the ratio was approximately equal to 1; for tapioca starch, the ratio was much lower than 1. Moreover, the starch granules of sweet potato and tapioca were both A-type crystallinity, while yam starch granules were C-type crystallinity. GrFFF could be used to characterize and separate large and small granules of tuber starches and estimate the ratio of starch granules A/B. Yam starch was suitable for the development of slowly digestible starch (one kind of functional food), which could delay the occurrence of metabolic syndrome, diabetes and cardiovascular diseases. Tapioca starch was suitable for plant capsules, which were safer than gelatin capsule and could prevent the "poison capsule" incident from happening. [ABSTRACT FROM AUTHOR]

The proto-oncogene Bcl3 induces survival and proliferation in cancer cells; however, its function and regulation in ovarian cancer (OC) remain unknown. Here, we show that Bcl3 expression is increased in human OC tissues. Surprisingly, however, we found that in addition to promoting survival, proliferation, and migration of OC cells, Bcl3 promotes both constitutive and interferon-γ (IFN)-induced expression of the immune checkpoint molecule PD-L1. The Bcl3 expression in OC cells is further increased by IFN, resulting in increased PD-L1 transcription. The mechanism consists of an IFN-induced, Bcl3- and p300-dependent PD-L1 promoter occupancy by Lys-314/315 acetylated p65 NF-kB. Blocking PD-L1 by neutralizing antibody reduces proliferation of OC cells overexpressing Bcl3, suggesting that the pro-proliferative effect of Bcl3 in OC cells is partly mediated by PD-L1. Together, this work identifies PD-L1 as a novel target of Bcl3, and links Bcl3 to IFNγ signaling and PD-L1-mediated immune escape. [ABSTRACT FROM AUTHOR]

Overexpression of the pro-angiogenic chemokine IL-8 (CXCL8) is associated with a poor prognosis in several solid tumors, including epithelial ovarian cancer (EOC). Even though histone deacetylase (HDAC) inhibition has shown remarkable antitumor activity in hematological malignancies, it has been less effective in solid tumors, including EOC. Here we report results that may explain the decreased efficiency of HDAC inhibition in EOC, based on our data demonstrating that HDAC inhibition specifically induces expression of IL-8/CXCL8 in SKOV3, CAOV3, andOVCAR3cells. Suppression or neutralization of vorinostat-induced IL-8/CXCL8 potentiates the vorinostat inhibitory effect on cell viability and proliferation. The IL-8/CXCL8 expression induced by vorinostat in EOC cells is dependent on IκB kinase (IKK) activity and associated with a gene-specific recruitment of IKKβ and IKK-dependent recruitment of p65 NFκB to the IL-8/CXCL8 promoter. In addition, HDACinhibition induces acetylation of p65 and histone H3 and their IL-8/CXCL8 promoter occupancy. In vivo results demonstrate that combining vorinostat and the IKK inhibitor Bay 117085 significantly reduces tumor growth in nude mice compared with control untreated mice or either drug alone. Mice in the combination group had the lowest IL-8/CXCL8 tumor levels and the lowest tumor expression of the murine neutrophil [7/4] antigen, indicating reduced neutrophil infiltration. Together, our results demonstrate that HDAC inhibition specifically induces IL-8/CXCL8 expression in EOC cells and that the mechanism involves IKK, suggesting that using IKK inhibitors may increase the effectiveness of HDAC inhibitors when treating ovarian cancer and other solid tumors characterized by increased IL-8/CXCL8 expression. [ABSTRACT FROM AUTHOR]

A sequential Diels-Alder reaction/rearrangement sequence was developed for the synthesis of diverse functionalized bicyclo[2.2.1]heptanes as novel floral and woody odorants. The outcome of the rearrangement depended on the substitution pattern of the dienes. 2D NMR analysis has established the correct relative configuration of the bicyclo[2.2.1]heptanone, which was originally misassigned. Furthermore, when the initiating DA reaction was catalyzed by a chiral Lewis acid, the bicyclo[2.2.1]heptane derivatives including (+)-herbanone can be obtained in an enantiomeric ratio (er) up to 96.5:3.5. [ABSTRACT FROM AUTHOR]

A common feature of progeria syndromes is a premature aging phenotype and an enhanced accumulation of DNA damage arising from a compromised repair system. HGPS (Hutchinson–Gilford progeria syndrome) is a severe form of progeria in which patients accumulate progerin, a mutant lamin A protein derived from a splicing variant of the lamin A/C gene (LMNA). Progerin causes chromatin perturbations which result in the formation of DSBs (double-strand breaks) and abnormal DDR (DNA-damage response). In the present article, we review recent findings which resolve some mechanistic details of how progerin may disrupt DDR pathways in HGPS cells. We propose that progerin accumulation results in disruption of functions of some replication and repair factors, causing the mislocalization of XPA (xeroderma pigmentosum group A) protein to the replication forks, replication fork stalling and, subsequently, DNA DSBs. The binding of XPA to the stalled forks excludes normal binding by repair proteins, leading to DSB accumulation, which activates ATM (ataxia telangiectasia mutated) and ATR (ATM- and Rad3-related) checkpoints, and arresting cell-cycle progression. [ABSTRACT FROM AUTHOR]

An unexpected gold(l)-catalyzed homo-Rautenstrauch rearrangement of 1-cyclopropyl propargylic esters to cydohexenones is disclosed. This rearrangement represents new evidence for the recently discussed gold-stabilized non-classical carbocation character of intermediates in gold catalysis. A mechanistic study proved partial chirality transfer from optically active propargyl acetates. [ABSTRACT FROM AUTHOR]

Human replication protein A (RPA), a heterotrimeric protein complex, was originally defined as a eukaryotic single‐stranded DNA binding (SSB) protein essential for the in vitro replication of simian virus 40 (SV40) DNA. Since then RPA has been found to be an indispensable player in almost all DNA metabolic pathways such as, but not limited to, DNA replication, DNA repair, recombination, cell cycle, and DNA damage checkpoints. Defects in these cellular reactions may lead to genome instability and, thus, the diseases with a high potential to evolve into cancer. This extensive involvement of RPA in various cellular activities implies a potential modulatory role for RPA in cellular responses to genotoxic insults. In support, RPA is hyperphosphorylated upon DNA damage or replication stress by checkpoint kinases including ataxia telangiectasia mutated (ATM), ATR (ATM and Rad3‐related), and DNA‐dependent protein kinase (DNA‐PK). The hyperphosphorylation may change the functions of RPA and, thus, the activities of individual pathways in which it is involved. Indeed, there is growing evidence that hyperphosphorylation alters RPA–DNA and RPA–protein interactions. In addition, recent advances in understanding the molecular basis of the stress‐induced modulation of RPA functions demonstrate that RPA undergoes a subtle structural change upon hyperphosphorylation, revealing a structure‐based modulatory mechanism. Furthermore, given the crucial roles of RPA in a broad range of cellular processes, targeting RPA to inhibit its specific functions, particularly in DNA replication and repair, may serve a valuable strategy for drug development towards better cancer treatment. J. Cell. Physiol. 208: 267–273, 2006. © 2006 Wiley‐Liss, Inc. [ABSTRACT FROM AUTHOR]

Using the fluid–magnetofluid formalism, we obtain axisymmetric stability criteria for a composite disc system consisting of stellar and gaseous magnetized singular isothermal discs (MSIDs). Both (M)SIDs are presumed to be razor thin and are gravitationally coupled in a self-consistent axisymmetric background equilibrium with power-law surface mass densities and flat rotation curves. The gaseous MSID is embedded with a non-force-free coplanar azimuthal magnetic field of radial scaling . Lou & Zou recently constructed exact global stationary configurations for both axisymmetric and non-axisymmetric coplanar magnetohydrodynamic (MHD) perturbations in such a composite MSID system and proposed the MHD Ds criteria for axisymmetric stability by the hydrodynamic analogy. In a different perspective, we derive and analyse here the time-dependent Wentzel–Kramers–Brillouin–Jeffreys (WKBJ) dispersion relation in the low-frequency and tight-winding regime to examine axisymmetric stability properties. By introducing a rotational Mach number Ds for the ratio of the stellar rotation speed Vs to the stellar velocity dispersion as, we readily determine the stable range of numerically to establish the Ds criteria for axisymmetric MSID stability. Those MSID systems rotating either too fast (ring fragmentation) or too slow (Jeans collapse) are unstable. The stable range of depends on three dimensionless parameters: the ratio λ for the Alfvén speed to the sound speed in the gaseous MSID, the ratio β for the square of the stellar velocity dispersion to the gas sound speed and the ratio δ for the surface mass densities of the two (M)SIDs. Our WKBJ results of (M)SID instability provide physically compelling explanations for the stationary analysis of Lou & Zou. We further introduce an effective MHD Q parameter for a composite MSID system and compare with the earlier work of Elmegreen, Jog and Shen & Lou. As expected, an axisymmetric dark matter halo enhances the stability against axisymmetric disturbances in a composite partial MSID system. In terms of the global star formation rate in a disc galaxy system, it would appear physically more sensible to examine the stability criterion against galactic observations. Relevance to large-scale structures in disc galaxies is also discussed. [ABSTRACT FROM AUTHOR]

We construct aligned and unaligned stationary perturbation configurations in a composite system of stellar and coplanarly magnetized gaseous singular isothermal discs (SIDs) coupled by gravity. This study extends recent analyses on (magnetized) SIDs by Shu et al., Lou and Lou & Shen. By this model, we intend to provide a conceptual framework to gain insights for multiwavelength large-scale structural observations of disc galaxies. Both SIDs are approximated to be razor thin and are in a self-consistent axisymmetric background equilibrium with power-law surface mass densities and flat rotation curves. The gaseous SID is embedded with a coplanar azimuthal magnetic field of a radial scaling that is not force-free. In comparison with the SID problems studied earlier, there are three possible classes of stationary solutions allowed by more dynamic freedoms. To identify physical solutions, we explore parameter space involving three dimensionless parameters: ratio λ of Alfvén speed to sound speed in the magnetized gaseous SID; ratio β of the square of the stellar velocity dispersion to the gas sound speed; and ratio δ of the surface mass densities of the two SIDs. For both aligned and unaligned spiral cases with azimuthal periodicities , one of the three solution branches is always physical, while the other two branches might become invalid when β exceeds certain critical values. For the onset criteria from an axisymmetric equilibrium to aligned secular bar-like instabilities, the corresponding ratio, which varies with λ, β and δ, may be considerably lower than the oft-quoted value of , where is the total kinetic energy, is the total gravitational potential energy and is the total magnetic energy. For unaligned spiral cases, we examine marginal instabilities for axisymmetric and non-axisymmetric disturbances. The resulting marginal stability curves differ from the previous ones. The case of a composite partial magnetized SID system is also investigated to include the gravitational effect of an axisymmetric dark matter halo on the SID equilibrium. We further examine the phase relationship among the mass densities of the two SIDs and azimuthal magnetic field perturbation. Our exact global perturbation solutions and critical points are valuable for testing numerical magnetohydrodynamic codes. For galactic applications, our model analysis contains more realistic elements and offers useful insights into the structures and dynamics of disc galaxies consisting of stars and magnetized gas. [ABSTRACT FROM AUTHOR]

During the DNA damage recognition of nucleotide excision repair in Escherichia coli the interaction of UvrB protein with damaged DNA ensures the recognition of differences in the intrinsic chemical structures of a variety of adduct molecules in DNA double helix. Our earlier study indicated that a single tyrosine-to-tryptophan mutation at residue 95 converted the UvrB to a protein [UvrB(Y95W)I that is able to bind to a structure-specific bubble DNA substrate, even in the absence of UvrA. fluorescence spectroscopy therefore was adopted to investigate the biochemical properties and thermodynamics of DNA damage recognition by the mutant protein. We examined the binding of the UvrB(Y95W) mutant protein to a structure-specific 30 bp DNA substrate containing a single fluorescein which serves as both an adduct and a fluorophore. Binding of the protein to the substrate results in a significant reduction in fluorescence. By monitoring the fluorescence changes, binding isotherms were generated from a series of titration experiments at various physiological temperatures, and dissociation constants were determined. Analysis of our data indicate that interaction of UvrB(Y95W) protein with the adduct incurred a large negative change in heat capacity ΔCp°obs (-1.1 kcal moI-1 K1), while the δG°obs was relatively unchanged with temperature. Further study of the binding at various concentrations of KCI showed that on average only about 1.5 ion pairs were involved in formation of the UvrB-DNA complex. Together, these results suggested that hydrophobic interactions are the main driving forces for the recognition of DNA damage by UvrB protein. [ABSTRACT FROM AUTHOR]

The DNA repair protein UvrB plays an indispensable role in the stepwise and sequential damage recognition of nucleotide excision repair in Escherichia coli. Our previous studies suggested that UvrB is responsible for the chemical damage recognition only upon a strand opening mediated by UvrA. Difficulties were encountered in studying the direct interaction of UvrB with adducts due to the presence of UvrA. We report herein that a single point mutation of Y95W in which a tyrosine is replaced by a tryptophan results in an UvrB mutant that is capable of efficiently binding to structure-specific DNA adducts even in the absence of UvrA. This mutant is fully functional in the UvrABC incisions. The dissociation constant for the mutant-DNA adduct interaction was less than 100 nM at physiological temperatures as determined by fluorescence spectroscopy. In contrast, similar substitutions at other residues in the β-hairpin with tryptophan or phenylalanine do not confer UvrB such binding ability. Homology modeling of the structure of E. coli UvrB shows that the aromatic ring of residue Y95 and only Y95 directly points into the DNA binding cleft. We have also examined UvrB recognition of both "normal" bulky BPDEDNA and protein-cross-linked DNA (DPC) adducts and the roles of aromatic residues of the β-hairpin in the recognition of these lesions. A mutation of Y92W resulted in an obvious decrease in the efficiency of UvrABC incisions of normal adducts, while the incision of the DPC adduct is dramatically increased. Our results suggest that Y92 may function differently with these two types of adducts, while the Y95 residue plays an unique role in stabilizing the interaction of UvrB with DNA damage, most likely by a hydrophobic stacking. [ABSTRACT FROM AUTHOR]

Examines the incision of DNA—protein crosslinks (DPC) by UvrABC nuclease. Recognition of biochemical pathways to repair lesions; Strategy to obtain site-specific DPC; Interaction of Escherichia coli on in vitro nucleotide excision repair.

This paper presents a comparison between CFD-simulations and measurements of the temperature stratification in a mixing box of an air-handling unit. We have used data from field measurements during a period of over a year for different outside temperatures. We performed two-dimensional CFD-simulations for four different outside temperatures with commercially available software. The measurements as well as the simulations show that the temperature difference between the upper part and the lower part of the duct downstream of the mixing box is considerable. It increases, as the outside temperature decreases. However, the discrepancies between the measurements and the simulations are large. The reasons for this are uncertain boundary conditions and modelling errors leading to an inaccurate simulation result. The stratification downstream of the mixing box implies large sensor errors and the use of the mixed air temperature for control and fault detection must therefore be questioned. Averaging sensors, which take a mean value over the duct section, can be used but do not consider differences in velocities and are therefore not accurate either. In order to, for example, use CFD as a tool to decide the optimal sensor location a more accurate model and more information regarding the boundary conditions is needed. Copyright © 2001 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Examines the hierarchy of DNA damage recognition in nucleotide excision repair in Escherichia coli. Differentiation between initial and second recognition; Role of adenosine triphosphate in DNA damage recognition; Factors influencing the incision efficiency of UvrABC.

Examines the differential incision of bulky carcinogen-DNA adducts by the UvrABC nuclease. Importance of Escherichia coli UvrABC nuclease system in the removal of DNA damages; Account on UvrABC proteins interaction in DNA lesion sites; Influence of different adducts on UvrA binding affinities; Extent of UvrB-DNA complex formation.

Repair proteins alter the local DNA structure during nucleotide excision repair (NER). However, the precise role of DNA melting remains unknown. A series of DNA substrates containing a unique site-specific BPDE-guanine adduct in a region of non-complementary bases were examined for incision by the Escherichia coli UvrBC endonuclease in the presence or absence of UvrA. UvrBC formed a pre-incision intermediate with a DNA substrate containing a 6-base bubble structure with 2 unpaired bases 5′ and 3 unpaired bases 3′ to the adduct. Formation of this bubble served as a dynamic recognition step in damage processing. UvrB or UvrBC may form one of three stable repair intermediates with DNA substrates, depending upon the state of the DNA surrounding the modified base. The dual incisions were strongly determined by the distance between the adduct and the double-stranded—single-stranded DNA junction of the bubble, and required homologous double-stranded DNA at both incision sites. Remarkably, in the absence of UvrA, UvrBC nuclease can make both 3′ and 5′ incisions on substrates with bubbles of 3–6 nucleotides, and an uncoupled 5′ incision on bubbles of i>10 nucleotides. These data support the hypothesis that the E.coli and human NER systems recognize and process DNA damage in a highly conserved manner. [ABSTRACT FROM AUTHOR]

In the presence of a Lewis acid, β,γ-unsaturated ketones and oximes or imines undergo nucleophilic addition to produce zwitterion intermediates, and subsequent oxy-2-azonia-Cope rearrangements give homoallylic amides. In the case of 2-vinylcycloalkanones, the process results in ring enlargement, providing a novel route to 9- to 16-membered lactams. The preparative significance of this protocol was evidenced by a short synthesis of macrocyclic alkaloid motuporamine G. The stereochemistry-defining step of this oxy-azonia-Cope rearrangement was further studied computationally. Despite a high-energy preequilibrium in the formation of zwitterionic intermediates, the [3,3]-sigmatropic step is the rate- and product-determining step. Chairlike transition states are generally preferred over boatlike ones. [ABSTRACT FROM AUTHOR]

We report a systematic and quantitative structure−function relationship study of the major N-deoxyguanosin-8-yl-2-aminofluorene adduct (AF) derived from the prototype carcinogen 2-aminofluorene and its derivatives. The AF adduct is known to exist in two distinct conformational motifs, depending upon the location of the hydrophobic fluorine moiety:  major groove binding “B type” (B) conformation (AF-dGanti) and base-displaced “stacked” (S) conformation (AF-dGsyn). The AF-induced S/B conformational heterogeneity is sequence-dependent and follows a typical two-site dynamic chemical exchange. The population of S conformation decreases in the order of 3‘-G > A C > T, indicating the importance of the purine flanking bases in promoting the stacking structure. Line-shape analysis showed that the S/B interconversion energy barriers (G) are in the narrow 14−16 kcal/mol range. The energy differences of the two conformers are relatively small (<0.5 kcal/mol), suggesting a possibility for a facile adduct conformation switch in the active site of a polymerase. The S/B equilibrium modulates the efficiency of Escherichia coliUvrABC-based nucleotide excision repair (NER) in a conformation-specific manner. The 19F NMR/NER results indicate greater repair susceptibility for the base-displaced S conformer, which lacks a Watson−Crick base pair at the lesion site. These findings represent the first of its kind quantitative structure−function work relating NER activity to a specific adduct conformer and will lead to a better understanding of how bulky DNA adducts are accommodated by the repair protein. [ABSTRACT FROM AUTHOR]

Replication protein A (RPA) is a eukaryotic single-stranded DNA-binding protein consisting of three subunits of 70-, 32-, and 14-kDa (RPA70, RPA32, RPA14, respectively). It is a protein essential for most cellular DNA metabolic pathways. Checkpoint proteins Rad9, Rad1, and Hus1 form a clamp-like complex which plays a central role in the DNA damage-induced checkpoint response. In this report, we presented the evidence that Rad9–Rad1–Hus1 (9–1–1) complex directly interacted with RPA in human cells, and this interaction was mediated by the binding of Rad9 protein to both RPA70 and RPA32 subunits. In addition, the cellular interaction of 9–1–1 with RPA or hyperphosphorylated RPA was stimulated by UV irradiation or camptothecin treatment in a dose-dependent manner. Such treatments also resulted in the colocalization of the nuclear foci formed with the two complexes. Consistently, knockdown of the RPA expression in cells by the small interference RNA (siRNA) blocked the DNA damage-dependent chromatin association of 9–1–1, and also inhibited the 9–1–1 complex formation. Taken together, our results suggest that 9–1–1 and RPA complexes collaboratively function in DNA damage responses, and that the RPA may serve as a regulator for the activity of 9–1–1 complex in the cellular checkpoint network.Oncogene (2005) 24, 4728–4735. doi:10.1038/sj.onc.1208674 Published online 9 May 2005 [ABSTRACT FROM AUTHOR]

Examines the intrastrand cross-links for the study of DNA replication and repair. Preparation of oligonucleotides containing intrastrand cross-links; Ability of oligonucleotides to bend DNA by Koo and Crothers method; Mobility of ligated oligomers.

Retinoblastoma is a childhood retinal tumor that develops from cone photoreceptor precursors in response to inactivating RB1 mutations and loss of functional RB protein. The cone precursor's response to RB loss involves cell type--specific signaling circuitry that helps to drive tumorigenesis. One component of the cone precursor circuitry, the thyroid hormone receptor β2 (TRβ2), enables the aberrant proliferation of diverse RB-deficient cells in part by opposing the down-regulation of S-phase kinase-associated protein 2 (SKP2) by the more widely expressed and tumor-suppressive TRβ1. However, it is unclear how TRβ2 opposes TRβ1 to enable SKP2 expression and cell proliferation. Here, we show that in human retinoblastoma cells TRβ2 mRNA encodes two TRβ2 protein isoforms: a predominantly cytoplasmic 54-kDa protein (TRβ2-54) corresponding to the well-characterized full-length murine Trβ2 and an N-terminally truncated and exclusively cytoplasmic 46-kDa protein (TRβ2-46) that starts at Met-79. Whereas TRβ2 knockdown decreased SKP2 expression and impaired retinoblastoma cell cycle progression, re-expression of TRβ2-46 but not TRβ2-54 stabilized SKP2 and restored proliferation to an extent similar to that of ectopic SKP2 restoration. We conclude that TRβ2-46 is an oncogenic thyroid hormone receptor isoform that promotes SKP2 expression and SKP2-dependent retinoblastoma cell proliferation. [ABSTRACT FROM AUTHOR]

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder that is caused by a point mutation in the LMNA gene, resulting in production of a truncated farnesylated-prelamin A protein (progerin). We previously reported that XPA mislocalized to the progerin-induced DNA double-strand break (DSB) sites, blocking DSB repair, which led to DSB accumulation, DNA damage responses, and early replication arrest in HGPS. In this study, the XPA mislocalization to DSBs occurred at stalled or collapsed replication forks, concurrent with a significant loss of PCNA at the forks, whereas PCNA efficiently bound to progerin. This PCNA sequestration likely exposed ds-ssDNA junctions at replication forks for XPA binding. Depletion of XPA or progerin each significantly restored PCNA at replication forks. Our results suggest that although PCNA is much more competitive than XPA in binding replication forks, PCNA sequestration by progerin may shift the equilibrium to favor XPA binding. Furthermore, we demonstrated that progerin-induced apoptosis could be rescued by XPA, suggesting that XPA-replication fork binding may prevent apoptosis in HGPS cells. Our results propose a mechanism for progerin-induced genome instability and accelerated replicative senescence in HGPS. [ABSTRACT FROM AUTHOR]

Fatty acid synthase (FASN), the sole cytosolic mammalian enzyme for de novo lipid synthesis, is crucial for cancer cell survival and associates with poor prognosis. FASN overexpression has been found to cause resistance to genotoxic insults. Here we tested the hypothesis that FASN regulates DNA repair to facilitate survival against genotoxic insults and found that FASN suppresses NF-κB but increases specificity protein 1 (SP1) expression. NF-κB and SP1 bind to a composite element in the poly(ADP-ribose) polymerase 1 (PARP-1) promoter in a mutually exclusive manner and regulate PARP-1 expression. Up-regulation of PARP-1 by FASN in turn increases Ku protein recruitment and DNA repair. Furthermore, lipid deprivation suppresses SP1 expression, which is able to be rescued by palmitate supplementation. However, lipid deprivation or palmitate supplementation has no effect on NF-κB expression. Thus, FASN may regulate NF-κB and SP1 expression using different mechanisms. Altogether, we conclude that FASN regulates cellular response against genotoxic insults by up-regulating PARP-1 and DNA repair via NF-κB and SP1. [ABSTRACT FROM AUTHOR]

XPA (xeroderma pigmentosum group A) protein is an essential factor for NER (nucleotide excision repair) which is believed to be involved in DNA damage recognition/verification, NER factor recruiting and stabilization of repair intermediates. Past studies on the structure of XPA have focused primarily on XPA interaction with damaged DNA. However, how XPA interacts with other DNA structures remains unknown though recent evidence suggest that these structures could be important for its roles in both NER and non-NER activities. Previously, we reported that XPA recognizes undamaged DNA ds/ssDNA (double-strand/single-strandDNA) junctions with a binding affinity much higher than its ability to bind bulky DNA damage. To understand how this interaction occurs biochemically we implemented a structural determination of the interaction using a MS-based protein footprinting method and limited proteolysis. By monitoring surface accessibility of XPA lysines to NHS-biotin modification in the free protein and the DNA junction-bound complex we show that XPA physically interacts with the DNA junctions via two lysines, K168 and K179, located in the previously known XPA(98-219) DBD (DNA-binding domain). Importantly, we also uncovered new lysine residues, outside of the known DBD, involved in the binding. We found that residues K221, K222, K224 and K236 in the C-terminal domain are involved in DNA binding. Limited proteolysis analysis of XPA-DNA interactions further confirmed this observation. Structural modelling with these data suggests a clamp-like DBD for the XPA binding to ds/ssDNA junctions. Our results provide a novel structure-function view of XPA-DNA junction interactions. [ABSTRACT FROM AUTHOR]

Cluster DNA damage refers to two or more lesions in a single turn of the DNA helix. Such clustering may occur with bulky DNA lesions, which may be responsible for their sequence-dependent repair and mutational outcomes. Here we prepared three 16-mer cluster duplexes in which two fluoroacetylaminofluorene adducts (dG-FAAF) are separated by zero, one, and two nucleotides in the Escherichia coli NarI mutational hot spot (5'-CTCTCG1G2CG3CCATCAC-3'): 5'-CG1*G2*CG3CC-3', 5'-CG1G2*CG3*CC-3', and 5'-CG1*G2CG3*CC-3' (G* = dG-FAAF), respectively. We conducted spectroscopic, thermodynamic, and molecular dynamics studies of these di-FAAF duplexes, and the results were compared with those of the corresponding mono-FAAF adducts in the same NarI sequence [Jain, V., et al. (2012) Nucleic Acids Res. 40, 3939-3951]. Our nucleotide excision repair results showed the diadducts were more reparable than the corresponding monoadducts. Moreover, we observed dramatic flanking base sequence effects on their repair efficiency in the following order: NarI-G2G3 > NarI-G1G3 > NarI-G1G2. The nuclear magnetic resonance, circular dichroism, ultraviolet melting, and molecular dynamics simulation results revealed that in contrast to the monoadducts, diadducts produced a synergistic effect on duplex destabilization. In addition, dG-FAAF at G2G3 and G1G3 destacks the neighboring bases, with greater destabilization occurring with the former. Overall, the results indicate the importance of base stacking and related thermal and thermodynamic destabilization in the repair of bulky cluster arylamine DNA adducts. [ABSTRACT FROM AUTHOR]

8,5'-Cyclopurines, making up an important class of ioniaing radiation-induced tandem DNA damage, are repaired only by nucleotide excision repair (NER). They accumulate in NER-impaired cells, as in Cockayne syndrome group B and certain Xeroderma Pigmentosum patients. A plasmid containing (S'S)-8,S'-cyclo-2'-deoxy- guanosine (S-cdG) was replicated in Eschericlsia coli with specific DNA polymerase knockouts. Viability was <1% in tbe wild-type strain, which increased to 5.3% with SOS. Viability decreased further in a pol II¯ strain, whereas it increased considerably in a pol IV sirain. Remarkably, no progenywas recovered from a pol V¯ strain, indicating that pol V is absolutely required for bypassing S-cdG. Progeny analyses indicated that S-cdG is significanily mutagenic, inducing ∼34% mutation with SOS. Most mutations were S-cdG → A mutations, though S-cdG → T mutation and deletion of 5'C also occurred. Incisions of purified Uv- rABC nuclease on S-cdG, S-cdA, and CS-dG-AP on a duplex St-mer showed that the incision rates are Cg-dG- AP > S-cdA> S-cdG. to summary, S-cdG is a major block to DNA replication, highly mutagenic, and repaired slowly in E. coli. [ABSTRACT FROM AUTHOR]

Abstract: An improved method for detecting early changes in tumors in response to treatment, based on a modification of diffusion-weighted magnetic resonance imaging, has been demonstrated in an animal model. Early detection of therapeutic response in tumors is important both clinically and in pre-clinical assessments of novel treatments. Noninvasive imaging methods that can detect and assess tumor response early in the course of treatment, and before frank changes in tumor morphology are evident, are of considerable interest as potential biomarkers of treatment efficacy. Diffusion-weighted magnetic resonance imaging is sensitive to changes in water diffusion rates in tissues that result from structural variations in the local cellular environment, but conventional methods mainly reflect changes in tissue cellularity and do not convey information specific to microstructural variations at sub-cellular scales. We implemented a modified imaging technique using oscillating gradients of the magnetic field for evaluating water diffusion rates over very short spatial scales that are more specific for detecting changes in intracellular structure that may precede changes in cellularity. Results from a study of orthotopic 9L gliomas in rat brains indicate that this method can detect changes as early as 24 h following treatment with 1,3-bis(2-chloroethyl)-1-nitrosourea, when conventional approaches do not find significant effects. These studies suggest that diffusion imaging using oscillating gradients may be used to obtain an earlier indication of treatment efficacy than previous magnetic resonance imaging methods. [Copyright &y& Elsevier]

Cellular accumulation of DNA damage has been widely implicated in cellular senescence, aging, and premature aging. In Hutchinson-Gilford progeria syndrome (HGPS) and restrictive dermopathy (RD), premature aging is linked to accumulation of DNA double-strand breaks (DSBs), which results in genome instability. However, how DSBs accumulate in cells despite the presence of intact DNA repair proteins remains unknown. Here we report that the recruitment of DSB repair factors Rad50 and Rad51 to the DSB sites, as marked by γ-H2AX, was impaired in human HGPS and Zmpste24-deficient cells. Consistently, the progeria-associated DSBs appeared to be uurepairable although DSBs induced by camptothecin were efficiently removed in the progeroid cells. We also found that these progeroid cells exhibited nuclear foci of xeroderma pigmentosum group A (XPA), a unique nucleotide excision repair protein. Strikingly, these XPA foci colocalized with the DSB sites in the progeroid cells. This XPA-DSB association was further confirmed and found to be mediated by DNA, using a modified chromatin immunoprecipitation assay and coimmunoprecipitation. RNA interference (RNAi) knockdown of XPA in HGPS cells partially restored DSB repair as evidenced by Western blot analysis, immunofluorescence and comet assays. We propose that the uncharacteristic localization of XPA to or near DSBs inhibits DSB repair, thereby contributing to the premature aging phenotypes observed in progeria arising from genetic defects in prelamin A maturation. [ABSTRACT FROM AUTHOR]

A systematic spectroscopic and computational study was conducted in order to probe the influence of base sequences on stacked (S) versus B-type (B) conformational heterogeneity induced by the major dG adduct derived from the model carcinogen 7-fluoro-2-aminofluorene (FAF). We prepared and characterized eight 12-mer DNA duplexes (-AG*N- series, d[CTTCTAG*NCCTC]; -CG*N- series, d[CTTCTCG*NCCTC]), in which the central guanines (G*) were site-specifically modified with FAF with varying flanking bases (N = G, A, C, T). S/B heterogeneity was examined by CD, UV, and dynamic 19F NMR spectroscopy. All the modified duplexes studied followed a typical dynamic exchange between the S and B conformers in a sequence dependent manner. Specifically, purine bases at the 3'-flanking site promoted the S conformation (G > A > C > T). Simulation analysis showed that the S/B energy barriers were in the 14-16 kcal/mol range. The correlation times (τ = I/K) were found to be in the millisecond range at 20 °C. The van der Waals energy force field calculations indicated the importance of the stacking interaction between the carcinogen and neighboring base pairs. Quantum mechanics calculations showed the existence of correlations between the total interaction energies (including electrostatic and solvation effects) and the S/B population ratios. The S/B equilibrium seems to modulate the efficiency of Escherichia coli UvrABC-based nucleotide excision repair in a conformation-specific manner: i.e., greater repair susceptibility for the S over B conformation and for the -AG*N- over the -CG*N- series. The results indicate a novel structure-function relationship, which provides insights into how bulky DNA adducts are accommodated by UvrABC proteins. [ABSTRACT FROM AUTHOR]

Human replication protein A (RPA), a heterotrimer composed of RPA70, RPA32, and RPA 14 subunits, contains four single-stranded DNA (ssDNA) binding domains (DBD): DBD-A, DBD-B, and DBD-C in RPA70 and DBD-D in RPA32. Although crystallographic or NMR structures of these DBDs and a trimerization core have been determined, the structure of the full length of RPA or the RPA-ssDNA complex remains unknown. In this article, we have examined the structural features of RPA interaction with ssDNA by fluorescence spectroscopy. Using a set of oligonucleotides (dT) with varying lengths as a molecular ruler and also as the substrate, we have determined at single-nucleotide resolution the relative positions of the ssDNA with interacting intrinsic tryptophans of RPA. Our results revealed that Trp528 in DBD-C and Trp 107 in DBD-D contact ssDNA at the 16th and 24th nucleotides (nt) from the 5′-end of the substrate, respectively. Evaluation of the relative spatial arrangement of RPA domains in the RPA-ssDNA complex suggested that DBD-B and DBD-C are spaced by about 4 nt (~19 Å) apart, whereas DBD-C and DBD-D are spaced by about 7 nt (~34 Å). On the basis of these geometric constraints, a global structure model for the binding of the major RPA DBDs to ssDNA was proposed. [ABSTRACT FROM AUTHOR]

The influence of DNA base sequence context on the removal of a bulky benzo[a]pyrene diol epoxide-guanine adduct, (+)-trans-B[a]P-N2-dG (G*), by UvrABC nuclease from the thermophilic organism Bacillus caldotenax was investigated. The lesion was flanked by either T or C in otherwise identical complementary 43-mer duplexes (TG*T or CG*C, respectively). It was reported earlier that in the CG*C context, a dominant minor groove adduct structure was observed by NMR methods with all Watson-Crick base pairs intact, and the duplex exhibited a rigid bend. In contrast, in the TG*T context, a highly flexible bend was observed, base pairing at G*, and two 5′-base pairs flanking the adduct were impaired, and multiple solvent-accessible adduct conformations were observed. The TG*T-43-mer duplexes are incised with consistently greater efficiency by UvrABC proteins from B. caldotenax by a factor of 2.3 ± 0.3. The rates of incisions increase with increasing temperature and are characterized by linear Arrhenius plots with activation energies of 27.0 ± 1.5 and 23.4 ± 1.0 kcal/mol for CG*C and TG*T duplexes, respectively. These values reflect the thermophilic characteristics of the UVrABC nuclease complex and the contributions of the different DNA substrates to the overall activation energies. These effects are consistent with base sequence context-dependent differences in structural disorder engendered by a loss of local base stacking interactions and Watson—Crick base pairing in the immediate vicinity of the lesions in the TG*T duplexes. The local weakening of base pairing interactions constitutes a recognition element of the UvrABC nucleotide excision repair apparatus. [ABSTRACT FROM AUTHOR]

Pierisin-1 is a potent apoptosis-inducing protein found in the pupal extract of the cabbage white butterfly. Pierisin-1 catalyzes the mono(ADP-ribosyl)ation of the 2‘-deoxyguanosine residue and produces a bulky adduct, N2-(ADP-ribos-1-yl)-2‘-deoxyguanosine (N2-ADPR-dG) in DNA. Here, we examined the involvement of the nucleotide excision repair (NER) system in the removal of N2-ADPR-dG in Escherichia coli(E. coli) and human cells. The results of mobility shift gel electrophoresis assays using a 50-mer oligodeoxynucleotide containing a single N2-ADPR-dG showed that E. coliUvrAB proteins bound to the N2-ADPR-dG in vitro. Incubation of the adducted oligodeoxynucleotides with UvrABC resulted in the incision of the oligonucleotides in vitro. The results of filter binding and gel mobility shift assays using human XPA protein showed that XPA bound to DNA containing N2-ADPR-dGs in vitro. Finally, we introduced plasmids containing N2-ADPR-dGs into E. coliand human cells. N2-ADPR-adducted plasmids replicated l0 times and 20 times less efficiently in NER-deficient E. coliand human cells than in their wild-type counterparts, respectively. More mutations were induced in the plasmid propagated in NER-deficient cells than that in wild-type human cells. These results indicate the involvement of the NER system in the repair of N2-ADPR-dG in both E. coliand human cells. [ABSTRACT FROM AUTHOR]

Human XPA is an important DNA damage recognition protein in nucleotide excision repair (NER). We previously observed that XPA binds to the DNA lesion as a homodimer [Liu, Y., Liu, Y., Yang, Z., Utzat, C., Wang, G., Basu, A. K., and Zou, Y. (2005) Biochemistry 44, 7361-7368]. Herein we report that XPA recognized undamaged DNA double-strand/single-strand (ds-ssDNA) junctions containing ssDNA branches with binding affinity (Kd = 49.1 ± 5.1 nM) much higher than its ability to bind to DNA damage. The recognized DNA junction structures include the Y-shape junction (with both 3′- and 5′-ssDNA branches), Y-overhang junction (with a 3′-ssDNA branch), and 5′-overhang junction (with a 5′-ssDNA branch). Using gel filtration chromatography and gel mobility shift assays, we showed that the highly efficient binding appeared to be carried out by the XPA monomer and that the binding was largely independent of RPA. Furthermore, XPA efficiently bound to six-nucleotide mismatched DNA bubble substrates with or without DNA adducts including C8 guanine adducts of AF, AAF, and AP and the T[6,4]T photoproducts. Using a set of defined DNA substrates with varying degrees of DNA bending, we also found that the XPC-HR23B complex recognized DNA bending, whereas neither XPA nor the XPA-RPA complex could bind to bent DNA. We propose that, besides DNA damage recognition, XPA may also play a novel role in stabilizing, via its high affinity to ds-ssDNA junctions, the DNA strand opening surrounding the lesion for stable formation of preincision NER intermediates. Our results provide a plausible mechanistic interpretation for the indispensable requirement of XPA for both global genome and transcription-coupled repairs. Since ds-ssDNA junctions are common intermediates in many DNA metabolic pathways, the additional potential role of XPA in cellular processes is discussed. [ABSTRACT FROM AUTHOR]

Nucleotide excision repair (NER) is a repair pathway that removes a variety of bulky DNA lesions in both prokaryotic and eukaryotic cells. The perturbation of DNA helix structure caused by the oxidative intrastrand lesions could render them good substrates for the NER pathway. Here we employed Escherichia coli NER enzymes, i.e., UvrA, UvrB, and UvrC, to examine the incision efficiency of duplex DNA carrying three different oxidative intrastrand cross-link lesions, that is, G[8–5]C, G[8–5m]mC, and G[8–5m]T, and two dithymine photoproducts, namely, the cis,syn-cyclobutane pyrimidine dimer (T[c,s]T) and the pyrimidine(6-4)pyrimidone product (T[6-4]T). Our results showed that T[6-4]T was the best substrate for UvrA binding, followed by G[8-5]C, G[8-Sm]mC, and G[8-Sm]T, and then by T[c,s]T. The efficiencies of the UvrABC incisions of these lesions were consistent with their UvrA binding affinities: the stronger the binding to UvrA, the higher the rate of incision. In addition, flanking DNA sequences appeared to have little effect on the binding affinity of UvrA for G[8-5]C as AG[8-5]CA was only slightly preferred over CG[8-5]CG. Consistently, these two sequences exhibited almost no difference in incision rates. Furthermore, we investigated the thermal stability of dodecameric duplexes containing G[8-5m]mC or G[8-5m]T, and our results revealed that these two lesions destabilized the duplex, due to an increase in the free energy for duplex formation at 37 °C, by approximately 5.4 and 3.6 kcal/mol, respectively. The destabilizations to the DNA helix caused by those lesions, for the most part, are correlated with the binding affinities of UvrA and incision rates of UvrABC. Taken together, the results from this study suggest that oxidative intrastrand lesions might be substrates for NER enzymes in vivo. [ABSTRACT FROM AUTHOR]

Human replication protein A (RPA), composed of RPA70, RPA32, and RPA14 subunits, undergoes hyperphosphorylation in cells in response to DNA damage. Hyperphosphorylation that occurs predominately in the N-terminal region of RPA32 is believed to play a role in modulating the cellular activities of RPA essential for almost all DNA metabolic pathways. To understand how the hyperphosphorylation modulates the functions of RPA, we compared the structural characteristics of full-length native and hyperphosphorylated RPAs using mass spectrometric protein footprinting, fluorescence spectroscopy, and limited proteolysis. Our mass spectrometric data showed that of 24 lysines and 18 arginines readily susceptible to small chemical reagent modification in native RPA, the three residues Lys-343, Arg-335, and Arg-382, located in DNA binding domain B (DBD-B) of RPA70, were significantly shielded in the hyperphosphorylated protein. Tryptophan fluorescence studies indicated significant quenching of Trp-361, located in the DBD-B domain, induced by hyperphosphorylation of RPA. Consistently, DBD-B became more resistant to the limited proteolysis by chymotrypsin after RPA hyperphosphorylation. Taken together, our results indicate that upon hyperphosphorylation of RPA32 N terminus (RPA32N), RPA undergoes a conformational change involving the single-stranded DNA binding cleft of DBD-B. Comparison of the interactions of native and hyperphosphorylated RPAs with short single-stranded oligonucleotides or partial DNA duplexes with a short 5′ or 3′ single-stranded DNA tails showed reduced affinity for the latter protein. We propose that the hyperphosphorylation may play a role in modulating the cellular pathways by altering the DBD-B-mediated RPA-DNA and RPA-protein interactions, hypothetically via the interaction of hyperphosphorylated RPA32N with DBD-B. [ABSTRACT FROM AUTHOR]

Human xeroderma pigmentosum group A (XPA) is an essential protein for nucleotide excision repair (NER). We have previously reported that XPA forms a homodimer in the absence of DNA. However, what oligomeric forms of XPA are involved in DNA damage recognition and how the interaction occurs in terms of biochemical understanding remain unclear. Using the homogeneous XPA protein purified from baculovirus-infected sf21 insect cells and the methods of gel mobility shift assays, gel filtration chromatography, and UV-cross-linking, we demonstrated that both monomeric and dimeric XPA bound to the DNA adduct of N-acetyl-2-aminofluorene (AAF), while showing little affinity for nondamaged DNA. The binding occurred in a sequential and protein concentration-dependent manner. At relatively low-protein concentrations, XPA formed a complex with DNA adduct as a monomer, while at the higher concentrations, an XPA dimer was involved in the specific binding. Results from fluorescence spectroscopic and competitive binding analyses indicated that the specific binding of XPA to the adduct was significantly facilitated and stabilized by the presence of the second XPA in a positive cooperative manner. This cooperative binding exhibited a Hill coefficient of 1.9 and the step binding constants of K1 = 1.4 × 106 M-1 and k2 = 1.8 × 107 M-1. When interaction of XPA and RPA with DNA was studied, even though binding of RPA-XPA complex to adducted DNA was observed, the presence of RPA had little effect on the overall binding efficiency. Our results suggest that the dominant form for XPA to efficiently bind to DNA damage is the XPA dimer. We hypothesized that the concentration-dependent formation of different types of XPA-damaged DNA complex may play a role in cellular regulation of XPA activity. [ABSTRACT FROM AUTHOR]

XPA plays an important role in the DNA damage recognition during human nucleotide excision repair. Here we report that the XPA is a homodimer either in the free state or as a complex with human RPA in solution under normal conditions. The human XPA protein purified from baculovirus-infected sf21 insect cells has a molecular mass of 36 317 Da, as determined by mass spectroscopy. However, the apparent molecular mass of XPA determined by the native gel filtration chromatography was about 71 kDa, suggesting that XPA is a dimer. This observation was supported by a native PFO--PAGE and fluorescence spectroscopy analysis. XPA formed a dimer (XPA[SUB2]) in a broad range of XPA and NaC1 concentrations, and the dimerization was not due to the disulfide bond formation. Furthermore, a titration analysis of the binding of XPA to the human RPA indicated that it was the XPA[SUB2] that formed the complex with RPA. Finally, the difference between the mass spectrometric and the calculated masses of XPA implies that the protein contains posttranslational modifications. Taken together, our data suggest that the dimerization of XPA may play an important role in the DNA damage recognition of nucleotide excision repair. [ABSTRACT FROM AUTHOR]

Provides information on a study which described concurrent measurement of tissue-specific fatty acid and glucose uptake in the conscious mouse undergoing rest or exercise with and without short-term inhibition of nitric oxide synthesis. Methodology of the study; Results and discussion on the study.

Hyperactivation of PARP1 is known to be a major cause of necrotic cell death by depleting NAD+/ATP pools during Ca2+ overload which is associated with many ischemic diseases. However, little is known about how PARP1 hyperactivity is regulated during calcium overload. In this study we show that ATR kinase, well known for its role in DNA damage responses, suppresses ionomycin, glutamate, or quinolinic acid-induced necrotic death of cells including SH-SY5Y neuronal cells. We found that the inhibition of necrosis requires the kinase activity of ATR. Specifically, ATR binds to and phosphorylates PARP1 at Ser179 after the ionophore treatments. This site-specific phosphorylation inactivates PARP1, inhibiting ionophore-induced necrosis. Strikingly, all of this occurs in the absence of detectable DNA damage and signaling up to 8 hours after ionophore treatment. Furthermore, little AIF was released from mitochondria/cytoplasm for nuclear import, supporting the necrotic type of cell death in the early period of the treatments. Our results reveal a novel ATR-mediated anti-necrotic mechanism in the cellular stress response to calcium influx without DNA damage signaling. [ABSTRACT FROM AUTHOR]

An abstract of the article "Structural and Thermodynamic Insight into Escherichia coli UvrABC-Mediated Incision of Cluster Diacetylaminofluorene Adducts on the NarI Sequence," by Vipin Jain and colleagues is presented.

This paper describes a tunable dual-wavelength Ti:sapphire laser system with quasi-continuous-wave and high-power outputs. In the design of the laser, it adopts a frequency-doubled Nd:YAG laser as the pumping source, and the birefringence filter as the tuning element. Tunable dual-wavelength outputs with one wavelength range from 700 nm to 756.5 nm, another from 830 nm to 900mn have been demonstrated. With a pump power of 23 W at 532 nm, a repetition rate of 7 kHz and a pulse width of 47.6 ns, an output power of 5.1 W at 744.8 nm and 860.9 nm with a pulse width of 13.2 ns and a line width of 3 nm has been obtained, it indicates an optical-to-optical conversion efficiency of 22.2%. [ABSTRACT FROM AUTHOR]

The genetic diseases Hutchinson-Gilford progeria syndrome (HGPS) and restrictive dermopathy (RD) arise from accumulation of farnesylated prelamin A because of defects in the lamin A maturation pathway. Both of these diseases exhibit symptoms that can be viewed as accelerated aging. The mechanism by which accumulation of farnesylated prelamin A leads to these accelerated aging phenotypes is not understood. Here we present evidence that in HGPS and RD fibroblasts, DNA damage checkpoints are persistently activated because of the compromise in genomic integrity. Inactivation of checkpoint kinases Ataxia-telangiectasia-mutated (ATM) and ATR (ATM- and Rad3-related) in these patient cells can partially overcome their early replication arrest. Treatment of patient cells with a protein farnesyltransferase inhibitor (FTI) did not result in reduction of DNA double-strand breaks and damage checkpoint signaling, although the treatment significantly reversed the aberrant shape of their nuclei. This suggests that DNA damage accumulation and aberrant nuclear morphology are independent phenotypes arising from prelamin A accumulation in these progeroid syndromes. Since DNA damage accumulation is an important contributor to the symptoms of HGPS, our results call into question the possibility of treatment of HGPS with FTIs alone. [ABSTRACT FROM AUTHOR]