共查询到20条相似文献,搜索用时 0 毫秒
1.
增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)是一种生长调控蛋白,在DNA复制、修复、细胞周期调控、基因外遗传(epigenetic inheritance)等事件的协同机制中发挥重要功能。PCNA的表达调控发生在多个层次,涉及ATFl、CREB、RFXl、p53、E2F等转录因子以及内含子指导的反义RNA等等。 相似文献
2.
Ricardo Sánchez Daniel Torres Jesús Prieto Francisco J. Blanco Ramón Campos-Olivas 《Biomolecular NMR assignments》2007,1(2):245-247
PCNA is an essential factor for DNA replication, repair, chromatin metabolism, and effector of cell-cycle regulatory signals.
The assignment of backbone 1HN, 13Cα, 13CO, and 15N, and sidechain 13Cβ resonances of the human PCNA homotrimeric ring (∼90 kDa, 261 residues) is reported here. 相似文献
3.
H W Chang Y C Lai C Y Cheng J L Ho S T Ding Y C Liu 《Journal of cellular biochemistry》1999,73(3):423-432
Proliferating cell nuclear antigen (PCNA), also known as a cofactor of DNA polymerase delta, is required for eukaryotic cell DNA synthesis and nucleotide excision repair. Expression of PCNA gene is growth-regulated and UV inducible. In our previous study, we have observed that the rat PCNA promoter has the serum responsiveness. In this study, we demonstrate its UV inducibility in CHO.K1 cells. The UV induction of the rat PCNA promoter activity was dose-dependent in the cells synchronized at different phases. In addition, the sequences of the promoter responsible for the UV inducibility were delimited to the region between nucleotides -70 and +125, which contains an AP-1 site and a downstream proximal ATF/CRE site. While mutation of the AP-1 site abrogated the UV inducibility, mutation of the ATF/CRE site enhanced the UV inducibility, suggesting that the two sites play different roles in the UV induction of the promoter. In addition, the role of p53 in the UV induction of rat PCNA promoter was investigated. We found that exogenous p53 was unable to mimic the UV irradiation to induce rat PCNA promoter and that the UV induction of the rat PCNA promoter was seen in p53 deficient cells. Therefore, it is unlikely that the UV induction of the rat PCNA promoter is p53 dependent. 相似文献
4.
Regulation of proliferating cell nuclear antigen during the cell cycle 总被引:53,自引:0,他引:53
The proliferating cell nuclear antigen (PCNA), also known as cyclin and DNA polymerase delta auxiliary factor, is present in reduced amounts in nongrowing cells and is synthesized at a greater rate in the S phase of growing cells. The recently discovered involvement of PCNA in DNA replication suggested that this pattern of expression functions to regulate DNA synthesis. We have investigated this possibility further by examining the synthesis, stability, and accumulation of PCNA in HeLa cells fractionated by centrifugal elutriation into nearly synchronous populations of cells at various positions in the cell cycle. In these fractionated cells we found that there is an increase in the rate of PCNA synthesis with a peak in early S phase of the cell cycle, but the magnitude of the increase is only 2-3-fold. This change reflects similar changes in the amount of PCNA mRNA. The fluctuating synthesis of PCNA maintains this protein at a roughly constant proportion of the total cell protein, although the amount doubles/cell in the cell cycle. Consistent with this observation, the stability of PCNA does not differ significantly from that of total cellular protein in synchronized HeLa cells. We also observed that a maximum of one-third of the total PCNA is tightly associated with the nucleus, presumably in replication complexes, at the peak of S phase. We conclude that the cyclic synthesis of PCNA in cycling HeLa cells maintains PCNA in excess of the amount involved directly in DNA replication and the amount of the protein neither fluctuates significantly with the cell cycle nor is limiting for DNA synthesis. 相似文献
5.
6.
7.
8.
Autoantibody to a nuclear antigen in proliferating cells. 总被引:85,自引:0,他引:85
9.
10.
Mechanism whereby proliferating cell nuclear antigen stimulates flap endonuclease 1 总被引:11,自引:0,他引:11
Human flap endonuclease 1 (FEN1), an essential DNA replication protein, cleaves substrates with unannealed 5'-tails. FEN1 apparently tracks along the flap from the 5'-end to the cleavage site. Proliferating cell nuclear antigen (PCNA) stimulates FEN1 cleavage 5-50-fold. To determine whether tracking, binding, or cleavage is enhanced by PCNA, we tested a variety of flap substrates. Similar levels of PCNA stimulation occur on both a cleavage-sensitive nicked substrate and a less sensitive gapped substrate. PCNA stimulates FEN1 irrespective of the flap length. Stimulation occurs on a pseudo-Y substrate that exhibits upstream primer-independent cleavage. A pseudo-Y substrate with a sequence requiring an upstream primer for cleavage was not activated by PCNA, suggesting that PCNA does not compensate for substrate features that inhibit cleavage. A biotin.streptavidin conjugation at the 5'-end of a flap structure prevents FEN1 loading. The addition of PCNA does not restore FEN1 activity. These results indicate that PCNA does not direct FEN1 to the cleavage site from solution. Kinetic analyses reveal that PCNA can lower the K(m) for FEN1 by 11-12-fold. Overall, our results indicate that after FEN1 tracks to the cleavage site, PCNA enhances FEN1 binding stability, allowing for greater cleavage efficiency. 相似文献
11.
Intranuclear localization of proliferating cell nuclear antigen during the cell cycle in renal cell carcinoma 总被引:2,自引:0,他引:2
Kawamura K Kobayashi Y Tanaka T Ikeda R Fujikawa-Yamamoto K Suzuki K 《Analytical and quantitative cytology and histology / the International Academy of Cytology [and] American Society of Cytology》2000,22(2):107-113
OBJECTIVE: To investigate, with laser scanning cytometry (LSC), proliferating cell nuclear antigen (PCNA) expression during the cell cycle in renal cell carcinoma. STUDY DESIGN: DNA ploidy and intracellular localization of PCNA in renal cell carcinoma were determined using LSC and immunohistochemistry. The subjects were nine patients who had received surgery for renal cell carcinoma. After DNA ploidy analysis, the glass slides were restained by immunohistochemistry of PCNA. LSC allowed direct observation of PCNA localization during the cell cycle because we could obtain immunohistochemical staining of PCNA as a function of cell cycle phase for individual cells. RESULTS: PCNA was not demonstrated in the nuclei of G0/G1 cells. PCNA expression increased from the S phase of the cell cycle. PCNA rapidly degraded at the end of the G2 phase. In the late G2 and M phase, PCNA was not detected in almost any nucleus. CONCLUSION: LSC allows morphologic observation of the intracellular distribution of PCNA during the cell cycle in renal cell carcinoma. 相似文献
12.
Structure of the human gene for the proliferating cell nuclear antigen 总被引:35,自引:0,他引:35
S Travali D H Ku M G Rizzo L Ottavio R Baserga B Calabretta 《The Journal of biological chemistry》1989,264(13):7466-7472
The proliferating cell nuclear antigen (PCNA, cyclin) was originally defined as a nuclear protein whose appearance correlated with the proliferative state of the cell. It is now known to be a co-factor of DNA polymerase delta and to be necessary for DNA synthesis and cell cycle progression. cDNA clones of human PCNA have been isolated and, using one of these cDNA, we have now obtained from a lambda phage library a clone containing the entire human PCNA gene and flanking sequences. The human PCNA gene is a unique copy gene and has 6 exons. It spans, from the cap site to the poly(A) signal 4961 base pairs. We have identified, in the 5'-flanking sequence, a region with promoter activity, a well as other structural elements common to other promoters. An interesting feature of the PCNA gene is the presence of extensive sequence similarities among introns and between introns and exons. 相似文献
13.
A cDNA fragment encoding a common bean (Phaseolus vulgaris) proliferating cell nuclear antigen (PCNA) was isolated using rapid amplification of cDNA 3' end (3' RACE) method, cloned and sequenced. The nucleotide sequence of this clone contains an open reading frame of 798 nucleotides encoding a protein of 265 amino acids. Alignment of the common bean PCNA predicted sequence shows its high degree of identity with PCNA from other plant species. Analysis of PCNA content in the germinating embryos of common bean showed a decrease in the protein level after 60h of germination. Moreover, PCNA was not detected in the tested plant organs (root, stem, leaf and flower). The presence of PCNA in the germinating seeds and its absence from mature plants suggests that this protein plays a crucial role during early stages of plant development. 相似文献
14.
D J Mozzherin C K Tan K M Downey P A Fisher 《The Journal of biological chemistry》1999,274(28):19862-19867
The relative positions of components of the DNA-dependent DNA polymerase delta (pol delta).proliferating cell nuclear antigen (PCNA).DNA complex were studied. We have shown that pol delta incorporates nucleotides close to a template biotin-streptavidin complex located 5' (downstream) to the replicating complex in the presence or absence of PCNA. PCNA-dependent synthesis catalyzed by pol delta was nearly totally (95%) inhibited by a biotin. streptavidin complex located at the 3'-end of a template with a 15-mer primer (upstream of the replicating complex), but was only partially inhibited with a 19-mer primer. With either primer, PCNA-independent synthesis was not affected by the biotin. streptavidin complex. Quantification of results with primers of varying length suggested that pol delta interacts with between 8 and 10 nucleotides of duplex DNA immediately proximal to the 3'-OH primer terminus. Using UV photocross-linking, we determined that the 125-kDa subunit of pol delta, but not the 50-kDa subunit, interacted with a photosensitive residue of a substrate oligonucleotide. Interaction apparently takes place through the C terminus of p125. Based on these results, we conclude that PCNA is located "behind" pol delta in the polymerization complex during DNA synthesis and that only the large subunit of pol delta (two-subunit form) interacts directly with DNA. A detailed model of the enzymatically active complex is proposed. 相似文献
15.
Uracil residues arise in DNA by the misincorporation of dUMP in place of dTMP during DNA replication or by the deamination of cytosine in DNA. Uracil-DNA glycosylase initiates DNA base excision repair of uracil residues by catalyzing the hydrolysis of the N-glycosylic bond linking the uracil base to deoxyribose. In human cells, the nuclear form of uracil-DNA glycosylase (UNG2) contains a conserved PCNA-binding motif located at the N-terminus that has been implicated experimentally in binding PCNA. Here we use purified preparations of UNG2 and PCNA to demonstrate that UNG2 physically associates with PCNA. UNG2 co-eluted with PCNA during size exclusion chromatography and bound to a PCNA affinity column. Association of UNG2 with PCNA was abolished by the addition of 100 mM NaCl, and significantly decreased in the presence of 10 mM MgCl(2). The functional significance of the UNG2.PCNA association was demonstrated by UNG2 activity assays. Addition of PCNA (30-810 pmol) to standard uracil-DNA glycosylase reactions containing linear [uracil-(3)H]DNA stimulated UNG2 catalytic activity up to 2.6-fold. UNG2 activity was also stimulated by 7.5 mM MgCl(2). The stimulatory effect of PCNA was increased by the addition of MgCl(2); however, the dependence on PCNA concentration was the same, indicating that the effects of MgCl(2) and PCNA on UNG2 activity occurred by independent mechanisms. Loading of PCNA onto the DNA substrate was required for stimulation, as the activity of UNG2 on circular DNA substrates was not affected by the addition of PCNA. Addition of replication factor C and ATP to reactions containing 90 pmol of PCNA resulted in two-fold stimulation of UNG2 activity on circular DNA. 相似文献
16.
Involvement of proliferating cell nuclear antigen (cyclin) in DNA replication in living cells. 总被引:20,自引:1,他引:19
下载免费PDF全文
![点击此处可从《Molecular and cellular biology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Proliferating cell nuclear antigen (PCNA) (also called cyclin) is known to stimulate the activity of DNA polymerase delta but not the other DNA polymerases in vitro. We injected a human autoimmune antibody against PCNA into unfertilized eggs of Xenopus laevis and examined the effects of this antibody on the replication of injected plasmid DNA as well as egg chromosomes. The anti-PCNA antibody inhibited plasmid replication by up to 67%, demonstrating that PCNA is involved in plasmid replication in living cells. This result further implies that DNA polymerase delta is necessary for plasmid replication in vivo. Anti-PCNA antibody alone did not block plasmid replication completely, but the residual replication was abolished by coinjection of a monoclonal antibody against DNA polymerase alpha. Anti-DNA polymerase alpha alone inhibited plasmid replication by 63%. Thus, DNA polymerase alpha is also required for plasmid replication in this system. In similar studies on the replication of egg chromosomes, the inhibition by anti-PCNA antibody was only 30%, while anti-DNA polymerase alpha antibody blocked 73% of replication. We concluded that the replication machineries of chromosomes and plasmid differ in their relative content of DNA polymerase delta. In addition, we obtained evidence through the use of phenylbutyl deoxyguanosine, an inhibitor of DNA polymerase alpha, that the structure of DNA polymerase alpha holoenzyme for chromosome replication is significantly different from that for plasmid replication. 相似文献
17.
Strzalka W Labecki P Bartnicki F Aggarwal C Rapala-Kozik M Tani C Tanaka K Gabrys H 《Journal of experimental botany》2012,63(8):2971-2983
Proliferating cell nuclear antigen (PCNA) is post-translationally modified in yeast and animal cells. Major studies carried out in the last decade have focused on the role of sumoylated and ubiquitinated PCNA. Using different approaches, an interaction between plant PCNA and SUMO both in vivo and in bacteria has been demonstrated for the first time. In addition, identical sumoylation patterns for both AtPCNA1 and 2 were observed in bacteria. The plant PCNA sumoylation pattern has been shown to differ significantly from that of Saccharomyces cerevisiae. This result contrasts with a common opinion based on previous structural analysis of yeast, human, and plant PCNAs, which treats PCNA as a highly conserved protein even between species. Analyses of AtPCNA post-translational modifications using different SUMO proteins (SUMO1, 2, 3, and 5) revealed similar modification patterns for each tested SUMO protein. Potential target lysine residues that might be sumoylated in vivo were identified on the basis of in bacteria AtPCNA mutational analyses. Taken together, these results clearly show that plant PCNA is post-translationally modified in bacteria and may be sumoylated in a plant cell at various sites. These data open up important new perspectives for further detailed studies on the role of PCNA sumoylation in plant cells. 相似文献
18.
Drosophila proliferating cell nuclear antigen. Structural and functional homology with its mammalian counterpart 总被引:9,自引:0,他引:9
L Ng G Prelich C W Anderson B Stillman P A Fisher 《The Journal of biological chemistry》1990,265(20):11948-11954
A protein with an apparent mass of 36 kDa was purified from Drosophila melanogaster embryos using a protocol developed for the purification of proliferating cell nuclear antigen (PCNA) from human 293 cells. The Drosophila protein comigrated with human PCNA on one-dimensional sodium dodecyl sulfate-polyacrylamide gels and cross-reacted with monoclonal anti-rabbit PCNA antibodies. NH2-terminal amino acid sequence analysis revealed that the putative Drosophila PCNA was highly homologous to human PCNA. Of the first 22 amino acids, 16 were identical, and 4 of the remaining 6 were changed conservatively. Results of total amino acid analysis were also consistent with a high degree of similarity between Drosophila PCNA and human PCNA. Functional analysis using the reconstituted simian virus 40 in vitro DNA replication system demonstrated that Drosophila PCNA could substitute, albeit with reduced efficiency, for human PCNA in stimulating simian virus 40 DNA synthesis. Affinity-purified anti-Drosophila PCNA antibodies cross-reacted with human PCNA and were able to recognize specifically Drosophila PCNA both on crude homogenate immunoblots and by indirect immunofluorescence analysis of proliferating cells in larval tissues in situ. These antibodies thus promise to be useful probes for the study of cell proliferation in this rapidly developing organism. 相似文献
19.
Human SHPRH suppresses genomic instability through proliferating cell nuclear antigen polyubiquitination
下载免费PDF全文
![点击此处可从《The Journal of cell biology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Motegi A Sood R Moinova H Markowitz SD Liu PP Myung K 《The Journal of cell biology》2006,175(5):703-708
Differential modifications of proliferating cell nuclear antigen (PCNA) determine DNA repair pathways at stalled replication forks. In yeast, PCNA monoubiquitination by the ubiquitin ligase (E3) yRad18 promotes translesion synthesis (TLS), whereas the lysine-63-linked polyubiquitination of PCNA by yRad5 (E3) promotes the error-free mode of bypass. The yRad5-dependent pathway is important to prevent genomic instability during replication, although its exact molecular mechanism is poorly understood. This mechanism has remained totally elusive in mammals because of the lack of apparent RAD5 homologues. We report that a putative tumor suppressor gene, SHPRH, is a human orthologue of yeast RAD5. SHPRH associates with PCNA, RAD18, and the ubiquitin-conjugating enzyme UBC13 (E2) and promotes methyl methanesulfonate (MMS)-induced PCNA polyubiquitination. The reduction of SHPRH by stable short hairpin RNA increases sensitivity to MMS and enhances genomic instability. Therefore, the yRad5/SHPRH-dependent pathway is a conserved and fundamental DNA repair mechanism that protects the genome from genotoxic stress. 相似文献
20.
Mammalian proliferating cell nuclear antigen stimulates the processivity of two wheat embryo DNA polymerases. 总被引:7,自引:2,他引:5
下载免费PDF全文
![点击此处可从《Plant physiology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Multiple DNA polymerases have been described in all organisms studied to date. Their specific functions are not easy to determine, except when powerful genetic and/or biochemical tools are available. However, the processivity of a DNA polymerase could reflect the physiological role of the enzyme. In this study, analogies between plant and animal DNA polymerases have been investigated by analyzing the size of the products synthesized by wheat DNA polymerases A, B, CI, and CII as a measure of their processivity. Thus, incubations have been carried out with poly(dA)-oligo(dT) as a template-primer under varying assay conditions. In the presence of MgCl2, DNA polymerase A was highly processive, whereas DNA polymerases B, CI, and CII synthesized much shorter products. With MnCl2 instead of MgCl2, DNA polymerase A was highly processive, DNA polymerases B and CII were moderately processive, and DNA polymerase CI remained strictly distributive. The effect of calf thymus proliferating cell nuclear antigen (PCNA) on wheat polymerases was studied as described for animal DNA polymerases. The high processivity of DNA polymerase A was PCNA independent, whereas both enzyme activity and processivity of wheat DNA polymerases B and CII were significantly stimulated by PCNA. On the other hand, DNA polymerase CI was not stimulated by PCNA and, like animal DNA polymerase beta, was distributive in all cases. From these results, we propose that wheat DNA polymerase A could correspond to a DNA polymerase alpha, DNA polymerases B and CII could correspond to the delta-like enzyme, and DNA polymerase CI could correspond to DNA polymerase beta. 相似文献