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1.
Acta Pharmaceutica Sinica B ; (6): 3406-3416, 2021.
Article in English | WPRIM | ID: wpr-922804

ABSTRACT

Non-small cell lung cancer is recognized as the deadliest cancer across the globe. In some areas, it is more common in women than even breast and cervical cancer. Its rise, vaulted by smoking habits and increasing air pollution, has garnered much attention and resource in the medical field. The first lung cancer treatments were developed more than half a century ago. Unfortunately, many of the earlier chemotherapies often did more harm than good, especially when they were used to treat genetically unsuitable patients. With the introduction of personalized medicine, physicians are increasingly aware of when, how, and in whom, to use certain anti-cancer agents. Drugs such as tyrosine kinase inhibitors, anaplastic lymphoma kinase inhibitors, and monoclonal antibodies possess limited utility because they target specific oncogenic mutations, but other drugs that target mechanisms universal to all cancers do not. In this review, we discuss many of these non-oncogene-targeting anti-cancer agents including DNA replication inhibitors (

2.
Article | IMSEAR | ID: sea-203803

ABSTRACT

There are multiple issues with setting up new diagnostic testing capacity outside ofregular diagnostic laboratories. This is not an overnight solution. Research laboratories havedifferent approaches, equipment and staff. There are multiple steps to ensure that themethods used and results are a correctly validated, as there is nothing more dangerous at themoment than reporting incorrect negative or positive results to staff or patients. That beingsaid we can adapt and validate assays and make them work using our equipment that can thenbe rolled out to support the NHS. PCR is a method widely used to rapidly make millions tobillions of copies of a specific DNA sample, allowing scientists to take a very small sample ofDNA and amplify it to a large enough amount to study in detail. It is fundamental too much ofgenetic testing including analysis of ancient samples of DNA and identification of infectiousagents. Using PCR, copies of very small amounts of DNA sequences are exponentially amplifiedin a series of cycles of temperature changes. PCR is now a common and often indispensabletechnique used in medical laboratory and clinical laboratory research for a broad variety ofapplications including biomedical research and criminal forensics.

3.
Journal of Medical Postgraduates ; (12): 808-813, 2020.
Article in Chinese | WPRIM | ID: wpr-823273

ABSTRACT

ObjectiveThere are few reports about abnormal oligonucleotide binding fold domain protein genes (OBGs) affecting the initiation of DNA replication in hepatocellular carcinoma through the microchromosome maintenance (MCM) complex. This study aims to explore the roles of reverse-transcription-related genes (RTGs) in Hepatocellular Carcinoma cells (HCC) and the correlation between gene polymorphisms and abnormal gene expression.Methods We created a mouse model by injecting hepatocellular carcinoma cell line H22 (logarithmic growth phase) and dissected the tumor bodies from tumor-forming mice. The control group was treated by isotonic saline without H22. The healthy liver tissue cells were taken from the control mice. The total RNA of the H22 group and control group were extracted, and differentially expressed genes were analyzed. Screening of differentially expressed reverse transcription-related DEGs (RDEGs), GO and KEGG analysis of RDEGs. The interaction analysis of RDEGs encoded proteins, and the correlation analysis of RDEGs polymorphism and gene expression.ResultsThere were 193 differentially expressed RTGs in HCCs, which were involved in two biological procedures, three cell components, one molecular function, three signal pathways, and three functional sites; Its function is mainly concentrated in DNA replication, especially the construction of MCM complex and telomere complex in which OBGs participate in the initiation of replication. Most related genes had OB fold domains. The results also showed that both AS and SNV caused gene polymorphism was positively correlated with gene expression, and most OBGs in HCC had SNV phenomenon, but not occurred in healthy liver tissue.Conclusion Collectively, AS and SNV may be important regulatory factors for gene expression. SNV may particularly affect the function of OBGs in the MCM complex to abnormally initiate DNA replication in HCC.

4.
Chinese Medical Journal ; (24): 2827-2835, 2018.
Article in English | WPRIM | ID: wpr-772915

ABSTRACT

Background@#DNA replication and sister chromatid cohesion 1 (DSCC1) (also called DCC1) is a component of an alternative replication factor C complex that loads proliferating cell nuclear antigen onto DNA during S phase of the cell cycle. It is located at 8q24 and frequently amplified in hepatocellular carcinoma (HCC). However, the role of DSCC1 in the carcinogenesis and progress of HCC has not been fully investigated. Here, we aimed to assert the importance of DSCC1 in the HCC.@*Methods@#In this study, copy number variation data and RNA sequencing data were used to calculate the DNA copy number and mRNA expression of DSCC1 in HCC. Quantitative polymerase chain reaction, Western blotting, and immunohistochemistry analysis were used to determine the mRNA and protein level of DSCC1 in HCC. The Kaplan-Meier analysis and univariate and multivariate Cox regression analysis were used to assess the association of DSCC1 with the overall survival (OS) of HCC patients. Moreover, lentiviral shRNA was used to knockdown DSCC1, and then, colony-forming assay, cell cycle assay, and cell proliferation assay were performed to evaluate the impact of DSCC1 silencing on HCC cell lines.@*Results@#We found that DSCC1 was amplified and highly expressed in HCC tumor tissues than in nontumor tissues. We then found that the overexpression of both mRNA and protein of DSCC1 was linked to the bad prognosis of HCC patients. Astonishingly, the protein level of DSCC1 was an independent prognostic factor for OS (hazard ratio, 1.79; 95% confidence interval, 1.17-2.74; P = 0.007). Furthermore, the clonogenic capacity of DSCC1-amplified HCC cell lines (MHCC-97H, MHCC-97L, and Hep3B) was significantly inhibited by transduction of a lentiviral shRNA that targets DSCC1. We also showed that knockdown of DSCC1 induced G0-G1 cell cycle arrest (increased from 60% to more than 80%) and greatly inhibited the proliferation of HCC cell lines.@*Conclusion@#These results suggest that DSCC1 is a putative HCC driver gene that promotes proliferation and is associated with poor prognosis in HCC.


Subject(s)
Female , Humans , Male , Middle Aged , Blotting, Western , Carcinoma, Hepatocellular , Genetics , Pathology , Cell Cycle , Genetics , Physiology , Cell Cycle Checkpoints , Genetics , Physiology , Cell Line, Tumor , Cell Proliferation , Genetics , Physiology , DNA Replication , Genetics , Physiology , Hep G2 Cells , Immunohistochemistry , Liver Neoplasms , Genetics , Pathology , Multivariate Analysis , Proportional Hazards Models , Real-Time Polymerase Chain Reaction
5.
Acta sci., Biol. sci ; 39(4): 463-467, Oct. - Dec. 2017. ilus
Article in English | LILACS | ID: biblio-877679

ABSTRACT

All living organisms need a DNA replication mechanism and it has been conserved in the three domains of life throughout evolutionary process. Primase is the enzyme responsible for synthesizing de novo RNA primers in DNA replication. Archaeo-Eukaryotic Primase (AEP) is the superfamily that typically forms a heterodimeric complex containing both a small catalytic subunit (PriS) and a large accessory noncatalytic subunit (PriL). Sulfolobus solfataricus is a model organism for research on the Genetics field. The aim of this work was to evaluate, via Bioinformatics tools, three mutations in the large subunit (PriL) of the archaeon Sulfolobus solfataricus. The aspartic acid residue in the positions (Asp) 62, (Asp) 235, (Asp) 241 have been substituted by glutamic acid (Glu). The highest positive free energy variation of the three substitutions analyzed occurred with the mutation at the (Asp) 241 site. The in silico analysis suggested that these mutations in PriL may destabilize its tridimensional structure interfering with replication mechanisms of Sulfolobus solfataricus. Moreover, it may also alter interactions with other molecules, making salt bridges, for instance.


Todos os organismos vivos necessitam de um eficiente mecanismo de replicação de DNA. Ao longo da evolução biológica foi observado que esse mecanismo é conservado nos três domínios da vida. Uma enzima importante que participa desse mecanismo é a RNA primase, a qual é responsável pela síntese de novo de iniciadores de RNA na replicação do DNA. Em Arquea-Eucariota, RNA Primase (AEP) tipicamente forma um complexo heterodimérico, que contém uma pequena subunidade catalítica (PriS) e uma subunidade maior não catalítica acessória (PriL). Sulfolobus solfataricus é um organismo modelo de Arquea para a pesquisa no campo da genética. O objetivo deste trabalho foi avaliar, por meio de ferramentas de bioinformática, três mutações pontuais na subunidade maior (PriL) de Sulfolobus solfataricus. Nas sequências mutantes, os resíduos de ácido aspártico nas posições (Asp) 62, (Asp) 235, (Asp) 241 foram substituídos por ácido glutâmico (Glu). A maior variação de energia livre positiva das três mutações analisadas ocorreu no sítio (Asp) 241. A análise in silico sugeriu que essas mutações em PriL podem desestabilizar sua estrutura tridimensional, interferindo com os mecanismos de replicação de Sulfolobus solfataricus. Além disso, podem alterar interações com outras moléculas, formando pontes salinas.


Subject(s)
Computer Simulation , DNA Replication Timing , Sulfolobus solfataricus , Mutation , RNA, Transfer, Met
6.
J Biosci ; 2016 June; 41(2): 173-182
Article in English | IMSEAR | ID: sea-181564

ABSTRACT

The interplay between the host and Human cytomegalovirus (HCMV) plays a pivotal role in the outcome of an infection. HCMV growth in endothelial and epithelial cells requires expression of viral proteins UL128, UL130, and UL131 proteins (UL128-131), of which UL130 is the largest gene and the only one that is not interrupted by introns.Mutation of the C terminus of the UL130 protein causes reduced tropism of endothelial cells (EC). However, very few host factors have been identified that interact with the UL130 protein. In this study, HCMV UL130 protein was shown to directly interact with the human protein Snapin in human embryonic kidney HEK293 cells by Yeast two-hybrid screening, in vitro glutathione S-transferase (GST) pull-down, and co-immunoprecipitation. Additionally, heterologous expression of protein UL130 revealed co-localization with Snapin in the cell membrane and cytoplasm of HEK293 cells using fluorescence confocal microscopy. Furthermore, decreasing the level of Snapin via specific small interfering RNAs decreased the number of viral DNA copies and titer inHCMV-infected U373-S cells. Taken together, these results suggest that Snapin, the pUL130 interacting protein, has a role in modulating HCMV DNA synthesis.

7.
Chinese Pharmacological Bulletin ; (12): 1199-1201, 2015.
Article in Chinese | WPRIM | ID: wpr-481832

ABSTRACT

MTH1 (MutT Homolog1 )as MutT homologous en-zyme,is a nucleotide pyrophosphatase,mainly involved in DNA damage repair process,especially plays an important role in the process of DNA replication in tumor cells.Recent studies have found that MTH1’s function is responsible for the development of a variety of tumors.Studies have shown that,MTH1 can remove tumor cells’oxidative DNA elements detrimental to the function-al structure,protecting tumor cell division and proliferation, maintaining tumor cell survival,however,normal cells do not need MTH1.Therefore,MTH1 may be only closely associated with abnormal cell growth.This makes MTH1 as therapeutic tar-gets that have been paid much attention.This article reviewed the relationship of MTH1 and tumor,discussed the mechanisms of MTH1 in tumor growth MTH1 and tumor treatment,so as to provide reference for clinical research and treatment of tumor.

8.
Article in Chinese | WPRIM | ID: wpr-850167

ABSTRACT

Objective To analyze the evolution of rtI233V mutation in the reverse transcriptase domain of hepatitis B virus (HBV) and its association with adefovir dipivoxil (ADV) resistance. Methods The rate of detection of rtI233V mutation in 9830 patients with chronic HBV infection was analyzed. HBV reverse transcriptase genes isolated from serial serum samples of two patients were amplified by nested PCR, and clonal sequencing (>20 clones/sample) was performed to analyze the evolution of rtI233V mutations. The replica of pTriEx-HBV1.1 vectors harboring wild-type and mutant strains (rtI233V, rtN236T, rtI233V+rtN236T) were respectively constructed and transiently transfected into HepG2 cells. Media containing serial concentrations of lamivudine (LAM), ADV, entecavir (ETV), or tenofovir (TDF) were used to treat the cells. Then HBV DNA in the supernatants was quantitatively determined by real-time PCR in order to analyze HBV mutants' replication competence and phenotypic characteristics under the drug pressures. Results The detection rate of rtI233V mutation in 9830 nucleos(t)ide analogues-treated patients was 0.28% (28/9830), including 0.19% (19 patients) with rtI233V individual mutation and 0.09% (9 patients) with rtI233V mutation combining with rtN236T or other mutations. All of the patients had rtI233V had ADV exposure history: 16 (57.1%) of them received ADV monotherapy for over six months, and 12(42.9%) of them received ADV combined sequential therapy for over 12 months. Replication competence and phenotypic resistance analysis showed rtI233V and wild-type strains had similar viral replication competence, while rtN236T exhibited significantly lower replication competence compared with wild-type strains. rtI233V strains remained sensitive to LAM, ADV, ETV, and TDF and showed little influence on drug resistance when combined with rtN236T, but it showed ability to restore the defected replication capacity of rtN236T strains. Conclusions The rtI233V mutation is associated with sub-optimal response to ADV. Though it does not directly reduce virus sensitivity to ADV, rtI233V mutation enhances replication competence of ADV resistant strains, and it is a complementary mutation.

9.
Genomics & Informatics ; : 136-144, 2014.
Article in English | WPRIM | ID: wpr-61851

ABSTRACT

Besides single-nucleotide variants in the human genome, large-scale genomic variants, such as copy number variations (CNVs), are being increasingly discovered as a genetic source of human diversity and the pathogenic factors of diseases. Recent experimental findings have shed light on the links between different genome architectures and CNV mutagenesis. In this review, we summarize various genomic features and discuss their contributions to CNV formation. Genomic repeats, including both low-copy and high-copy repeats, play important roles in CNV instability, which was initially known as DNA recombination events. Furthermore, it has been found that human genomic repeats can also induce DNA replication errors and consequently result in CNV mutations. Some recent studies showed that DNA replication timing, which reflects the high-order information of genomic organization, is involved in human CNV mutations. Our review highlights that genome architecture, from DNA sequence to high-order genomic organization, is an important molecular factor in CNV mutagenesis and human genomic instability.


Subject(s)
Humans , Base Sequence , DNA , DNA Copy Number Variations , DNA Replication , DNA Replication Timing , Genome , Genome, Human , Genomic Instability , Mutagenesis , Recombination, Genetic
10.
Article in Chinese | WPRIM | ID: wpr-850403

ABSTRACT

Objective:To evaluate the influence of hepatitis B virus (HBV) genome nucleotide A1846T mutation on the viral replication capacity and the transcription activity of HBV core promoter (CP) in vitro. Method A total of 385 patients with hepatitis B admitted to the 302 Hospital of PLA were enrolled in the study, including 116 with moderate chronic hepatitis B (CHB-M), 123 with severe chronic hepatitis B (CHB-S), and 146 with acute-on-chronic liver failure (ACLF). Serum HBV DNA was isolated and full-length HBV genome was amplified. The incidence of A1846T was analyzed. Full-length HBV genomes containing 1846T mutation were cloned into pGEM-T easy vector, and the counterpart wild-type 1846A plasmids were obtained by sitedirected mutagenesis. The full-length HBV genome was released from recombinant plasmid by BspQ /Sca digestion, and then transfected into HepG2 cells. Secreted HBsAg level and intracellular HBV core particles were measured 72 hours post-transfection to analyze the replication capacity (a 1.0-fold HBV genome model). 1846 mutant and wild-type full-length HBV genomes were extracted to amplify the fragment of HBV CP region, and the dual luciferase reporter of the pGL3-CP was constructed. The luciferase activity was detected 48 hours post-transfection. Result The incidence of A1846T mutation gradually increased with the severity of hepatitis B, reaching 31.03%, 42.27%, and 55.48% in CHB-M, CHB-S and ACLF patients respectively (P0.01). The replication capacity of 1846T mutants, level of secreted HBsAg, and transcriptional activity of CP promoter were increased by 320%, 28% and 85% respectively, compared with 1846A wild-type strains. While the more common double mutation A1762T/ G1764A in CP region was increased by 67%, 9% and 72% respectively, compared with its counterpart wild-type strains. A1846T had a greater influence on viral replication capacity in vitro. Conclusions A1846T mutation could significantly increase the replication capacity of hepatitis B virus, secretion of HBsAg and transcription activity of CP promoter, and cis-activate the downstream gene transcription. The finding indicates that HBV genome A1846T mutation might play a role in liver disease progression.

11.
Article in Korean | WPRIM | ID: wpr-43862

ABSTRACT

DNA replication of human cytomegalovirus (HCMV) is a highly regulated process that requires specific interactions between cis-acting lytic origin of replication (oriLyt) and trans-acting viral proteins. Formation of the replication initiation complex is also regulated by specific interactions among viral replication proteins. HCMV replication proteins include origin-binding proteins, core proteins that work in replication forks, and regulatory proteins that modulate host cell functions. This letter describes intriguing questions regarding how HCMV origin-binding proteins interact with oriLyt to initiate DNA replication and how the regulatory UL112-113 proteins, which are found only in beta-herpesviruses, function to promote viral DNA replication.


Subject(s)
Humans , Cytomegalovirus , DNA , DNA Replication , DNA, Viral , Proteins , Replication Origin , Viral Proteins
12.
Article in English | WPRIM | ID: wpr-192552

ABSTRACT

Human bone marrow mesenchymal stem cells (MSCs) expanded in vitro exhibit not only a tendency to lose their proliferative potential, homing ability and telomere length but also genetic or epigenetic modifications, resulting in senescence. We compared differential methylation patterns of genes and miRNAs between early-passage [passage 5 (P5)] and late-passage (P15) cells and estimated the relationship between senescence and DNA methylation patterns. When we examined hypermethylated genes (methylation peak > or = 2) at P5 or P15, 2,739 genes, including those related to fructose and mannose metabolism and calcium signaling pathways, and 2,587 genes, including those related to DNA replication, cell cycle and the PPAR signaling pathway, were hypermethylated at P5 and P15, respectively. There was common hypermethylation of 1,205 genes at both P5 and P15. In addition, genes that were hypermethylated at P5 (CPEB1, GMPPA, CDKN1A, TBX2, SMAD9 and MCM2) showed lower mRNA expression than did those hypermethylated at P15, whereas genes that were hypermethylated at P15 (MAML2, FEN1 and CDK4) showed lower mRNA expression than did those that were hypermethylated at P5, demonstrating that hypermethylation at DNA promoter regions inhibited gene expression and that hypomethylation increased gene expression. In the case of hypermethylation on miRNA, 27 miRNAs were hypermethylated at P5, whereas 44 miRNAs were hypermethylated at P15. These results show that hypermethylation increases at genes related to DNA replication, cell cycle and adipogenic differentiation due to long-term culture, which may in part affect MSC senescence.


Subject(s)
Humans , Bone Marrow Cells/metabolism , DNA Methylation , Gene Expression Profiling , Gene Expression Regulation , Mesenchymal Stem Cells/metabolism , MicroRNAs , Molecular Sequence Annotation , Primary Cell Culture , Promoter Regions, Genetic , Reproducibility of Results , Signal Transduction , Telomere Shortening
13.
Article in Chinese | WPRIM | ID: wpr-391396

ABSTRACT

Objective To construct recombinant full length genotype B and C hepatitis B virus (HBV)and to examine HBV DNA replication and hepatitis B surface antigen(HBsAg),hepatitis B e antigen(HBeAg)expressions in Huh7 cells. Methods The full length genotype B and C HBV DNA were extracted and amplified from two HBV infected patients. The recombinant plasmids were constructed by inserting the amplified HBV fragments into the eukaryotic expression vector,pHY106,which were then transfected into Huh7 cells. The cells transfected with blank pHY106 vector were used as control. HBV DNA replication at 72 hours of transfection was detected by Southern blot. The HBV DNA levels in Huh7 cells at 24,48,72,96 and 120 hours of transfection were determined by real-time polymerase chain reaction(PCR).Meanwhile, the HBsAg and HBeAg expression levels in the supernatants at 24,48,72,96 and 120 hours were determined by enzyme linked immunosorbent assay(ELISA).Results The recombinant plasmids expressing genotype B or C HBV DNA were successfully constructed.The HBV replicative intermediates in HBV core particles,including rcDNA dsDNA and ssDNA,were detected by Southern blot.HBV DNA level could reach 8 lg copy/mL which was by real-time PCR. HBsAg and HBeAg levels determined by ELISA peaked at 72 hours after transfection and then declined gradually. Conclusions The recombinant plasmids inserted with genotype B or C HBV DNA are constructed successfully, which can express high levels of HBsAg and HBeAg in Huh7 cells. This system provides a platform for studying the pathogenesis of B and C genotype HBV, the interaction between HBV and host, as well as exploiting new drugs against HBV.

14.
Article in Korean | WPRIM | ID: wpr-98600

ABSTRACT

PURPOSE: Reactivation of hepatitis B virus (HBV) replication after hepatic resection might be a significant risk factor for prognosis in patients with chronic hepatitis B. The purpose of the present study was to investigate the changing pattern of serum HBV DNA titer after hepatic resection and to assess the incidence of reactivation of HBV replication. METHODS: Among HBV-positive patients who underwent hepatic resection for hepatocellular carcinoma, thirty-six patients with preoperative serum HBV DNA titer > or =3 log(10)copies/mL were enrolled. Serum DNA titers were examined before the operation, on the second and seventh postoperative days, and one month after the operation. RESULTS: The serum DNA titer decreased on the second postoperative day (p=0.078). The DNA level, however, had substantially returned to preoperative values by the seventh postoperative day (p<0.001). For most patients, the postoperative DNA titer reached its zenith on the seventh postoperative day or one month after the operation. The zenith level was higher (by 0.49+/-0.25 log10copies/mL) than preoperative levels although this difference just missed significance (p=0.068). Although postoperative reactivation of HBV replication emerged in 6 patients, only one of those patients developed postoperative hepatitis. Overall, four patients developed postoperative hepatitis and all of them had high postoperative HBV DNA levels (over 6 log(10)copies/mL). CONCLUSION: Although serum HBV DNA titers tended to increase postoperatively, routine antiviral therapy might be unnecessary because of the low incidence of postoperative hepatitis. High postoperative DNA levels, however, might be a risk factor for hepatitis, and postoperative follow-up of serum HBV DNA levels might be necessary in HBV-positive patients with hepatic resection.


Subject(s)
Humans , Carcinoma, Hepatocellular , DNA , DNA Replication , Follow-Up Studies , Hepatectomy , Hepatitis , Hepatitis B , Hepatitis B virus , Hepatitis B, Chronic , Incidence , Prognosis , Risk Factors
15.
Virologica Sinica ; (6): 243-267, 2009.
Article in Chinese | WPRIM | ID: wpr-671425

ABSTRACT

Baculoviruses were first identified as insect-specific pathogens, and it was this specificity that lead to their use as safe, target specific biological pesticides. For the past 30 years, AcMNPV has served as the subject of intense basic molecular research into the baculovirus infectious cycle including the interaction of the virus with a continuous insect cell line derived from Spodoptera frugiperda. The studies on baculoviruese have led to an in-depth understanding of the physical organization of the viral genomes including many complete genomic sequences, the time course of gene expression, and the application of this basic research to the use of baculoviruses not only as insecticides, but also as a universal eukaryotic protein expression system, and a potential vector in gene therapy. A great deal has also been discovered about the viral genes required for the replication of the baculovirus genome, while much remains to be learned about the mechanism of viral DNA replication. This report outlines the current knowledge of the factors involved in baculovirus DNA replication, using data on AcMNPV as a model for most members of the Baculoviridae.

16.
Genet. mol. res. (Online) ; 7(1): 127-132, Jan. 2008. ilus, tab
Article in English | LILACS | ID: lil-553779

ABSTRACT

DNA damage activates several mechanisms such as DNA repair and cell cycle checkpoints. The Saccharomyces cerevisiae heterotrimeric checkpoint clamp consisting of the Rad17, Mec3 and Ddc1 subunits is an early response factor to DNA damage and activates checkpoints. This complex is structurally similar to the proliferating cell nuclear antigen (PCNA), which serves as a sliding clamp platform for DNA replication. Growing evidence suggests that PCNA-like complexes play a major role in DNA repair as they have been shown to interact with and stimulate several proteins, including specialized DNA polymerases. With the aim of extending our knowledge concerning the link between checkpoint activation and DNA repair, we tested the possibility of a functional interaction between the Rad17/Mec3/Ddc1 complex and the replicative DNA polymerases alpha, delta and epsilon. The analysis of sensitivity response of single and double mutants to UVC and 8-MOP + UVA-induced DNA damage suggests that the PCNA-like component Mec3p of S. cerevisiae neither relies on nor competes with the third subunit of DNA polymerase delta, Pol32p, for lesion removal. No enhanced sensitivity was observed when inactivating components of DNA polymerases alpha and epsilon in the absence of Mec3p. The hypersensitivity of pol32delta to photoactivated 8-MOP suggests that the replicative DNA polymerase delta also participates in the repair of mono- and bi-functional DNA adducts. Repair of UVC and 8-MOP + UVA-induced DNA damage via polymerase delta thus occurs independent of the Rad17/Mec3/Ddc1 checkpoint clamp.


Subject(s)
Cell Cycle Proteins , DNA-Directed DNA Polymerase/metabolism , DNA Repair , Phosphoproteins/metabolism , Nuclear Proteins/metabolism , DNA-Binding Proteins/metabolism , Saccharomyces cerevisiae/enzymology , Proliferating Cell Nuclear Antigen/metabolism , DNA-Directed DNA Polymerase/classification , DNA, Fungal , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/genetics
17.
Virologica Sinica ; (4): 94-107, 2007.
Article in Chinese | WPRIM | ID: wpr-635185

ABSTRACT

Baculoviruses are the only nuclear replicating DNA-containing viruses that encode their own DNA-directed RNA polymerase (RNAP). The baculovirus RNAP is specific for the transcription of genes expressed after virus DNA replication. It is composed of four subunits, making it the simplest multisubunit RNAP known. Two subunits contain motifs found at the catalytic center of other RNAPs and a third has capping enzyme functions. The function of the fourth subunit is not known. Structural studies on this unique RNAP will provide new insights into the functions of this enzyme and the regulation of viral genes and may be instrumental to optimize the baculovirus gene expression system.

18.
Article in Chinese | WPRIM | ID: wpr-592162

ABSTRACT

The investigations on the interactive relationships of DNA replication, damage repair and recombination have been locating in the frontline and becoming one of the hotspots in today's life science research. More and more studies show that the processes of DNA replication, damage repair and recombination are both independent and interdependent in the molecular level. These pathways coordinate and conform each other through interactions of many critical proteins in the pathways, by which DNA molecules, known as genetical materials, can be well maintained in cell and faithfully transferred through cellular generations. By using E. coli as a model system, the recent progresses and the possible rules underlying E. coli DNA replication, repair and recombination have been analyzed. As it is believed that the researches on E. coli DNA replication, repair and recombination may be capable of providing clues to the eukaryotic research based on the universal conservations of the critical proteins in the pathways.

19.
Article in Chinese | WPRIM | ID: wpr-586055

ABSTRACT

Cell cycle checkpoints are protective mechanism responding to DNA damages originated from externalor internal factors. When cells are exposed to genotoxic stress or when nutrition crisis occurs, cell cycleprogression is usually stopped or slowed down by cell cycle checkpoints to allow for DNA repair or for handlingthe crisis. Besides, recent studies suggest that some cell cycle checkpoint proteins are also involved in regulatingphysiological DNA replication via controlling the rate of DNA replication. Cell cycle checkpoint proteins ATR,9-1-1 complex, Chk1, Cdc25A and CDK2 may participate in this process. This kind of regulation is supposed to bevery important for ensuring accurate DNA replication and maintaining genomic stability.

20.
Article in Chinese | WPRIM | ID: wpr-323011

ABSTRACT

This study examined the effect of artesunate (Art) on the proliferation, DNA replication, cell cycles and apoptosis of vascular smooth muscle cells (VSMCs). Primary cultures of VSMCs were established from aortas of mice and artesunate of different concentrations was added into the medium. The number of VSMCs was counted and the curve of cell growth was recorded.The activity of VSMCs was assessed by using MTT method and inhibitory rate was calculated.DNA replication was evaluated by [3 H]-TdR method and apoptosis by DNA laddering and HE staining. Flowmetry was used for simultaneous analysis of cell apoptosis and cell cycles. Compared with the control group, VSMCs proliferation in Art interfering groups were inhibited and [3H]-TdR incorprating rate were decreased as well as cell apoptosis was induced. The progress of cell cycle was blocked in G0/G1 by Art in a dose-dependent manner. It is concluded that Art inhibits VSMCs proliferation by disturbing DNA replication, inducing cell apoptosis and blocking cell cycle in G0/G1 phase.

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