Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.7 (DNA polymerase)
17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Aphidicolin, a specific and direct inhibitor of eukaryotic DNA polymerase alpha, was used to investigate its impact on immunologic reactions in vitro. Dose response curve of the inhibitory effect was studied in murine and human primary allogeneic responses, as well as the proliferative responses to both PHA and Con A mitogens. The presence of aphidicolin during the allosensitization phase in secondary MLR of mice splenocytes resulted in complete abolishment of the subsequent response directed against the priming alloantigens, whereas alloreactivity to unrelated alloantigen-bearing cells was inhibited to a much lesser degree. The allosensitized aphidicolin-treated cells lost the ability to respond to subsequent PHA stimulation, but were capable of exerting a high responsiveness to Con A. The presence of aphidicolin during the allosensitization phase in secondary MLR of human mononuclear cells resulted in markedly decreased alloreactivity directed against the priming cells, but spared the subsequent response to unrelated alloantigens and to both PHA and Con A mitogenic stimuli. It is suggested that aphidicolin may be used for selective inactivation of proliferating cells without interfering with immunologic functions of other quiescent subsets. Aphidicolin may thus be a useful agent for induction of specific unresponsiveness in experimental models of allogeneic transplantation.
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PMID:Selective abrogation of alloreactivity via priming in the presence of aphidicolin, a specific inhibitor of DNA polymerase. 252 76

The cell cycle dependent fluctuation of adenosine diphosphoribosyl transferase (ADPRT) activity was demonstrated by both nicotinamide adenine dinucleotide (3H-NAD+) incorporation into the acid insoluble fraction of permeabilized cells and changes in the cellular content of NAD, the only substrate of ADPRT, in intact FL cells. The ADPRT activity was lowest in the G1 phase and highest in the S/G2-G2 phase. Aphidicolin, a specific inhibitor of DNA polymerase a, abolished the fluctuation of ADPRT activity. Meanwhile, in 5-fluorodeoxy-uridine (FUdR) exposed cells whose DNA synthesis was interfered with by the inhibition of thymidylate synthetase and the rate of ligation of short replicative intermediates, the ADPRT activity remained at a higher level than in controls. However, 3-aminobenzamide (3AB), a potent ADPRT inhibitor, showed down DNA synthesis in the S phase and also extended the S phase. These results indicate that ADP-ribosylation may be involved in DNA replication and cell cycle progression, and suggest that ADPRT activity may be stimulated by transient short fragments of newly replicated DNA, exerting its effects at the later stages of DNA replication, most probably at the ligation step of DNA synthesis.
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PMID:On the relationship between adenosine diphosphoribosyl transferase and S phase DNA synthesis in cultured mammalian cells. 253 93

Epstein-Barr virus (EBV)-specified DNA polymerase was purified from P3HR-1 cells, a Burkitt lymphoma EBV producer cell line, treated with phorbol-12,13-dibutyrate (PDB) and n-butyrate. Its inhibition by aphidicolin, phosphonoformate (PFA) and 5'-GMP was examined. Aphidicolin could inhibit EBV DNA polymerase competitively with respect to dATP and dCTP and noncompetitively with respect to dGTP and dTTP; whereas 5'-GMP was a noncompetitive inhibitor with respect to all four dNTPs. Combinations of aphidicolin and PFA, or PFA and 5'-GMP, produced a mutually exclusive inhibition pattern of EBV DNA polymerase that suggested that the binding sites of these compounds on the enzyme molecule are kinetically overlapping.
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PMID:Interaction of Epstein-Barr virus DNA polymerase with aphidicolin, phosphonoformate and 5'-GMP. 285 12

Aphidicolin, a specific inhibitor of cellular DNA polymerase alpha and of viral DNA polymerase, inhibits production of infectious virus and cellular and viral DNA synthesis of human cytomegalovirus (HCMV)-infected cells. On the other hand, 2',3'-dideoxynucleosides, inhibitors of DNA polymerases beta and gamma, do not affect HCMV replication. The data show that the alpha DNA polymerases of either viral or cellular origin are required for viral DNA synthesis, and cannot be substituted by the cellular DNA polymerase beta and gamma.
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PMID:Effect of DNA polymerase inhibitors on the replication of human cytomegalovirus. Brief report. 298 30

Aphidicolin, a specific inhibitor of DNA polymerase alpha, provided a novel method for distinguishing between initiation of DNA synthesis at the simian virus 40 (SV40) origin of replication (ori) and continuation of replication beyond ori. In the presence of sufficient aphidicolin to inhibit total DNA synthesis by 50%, initiation of DNA replication in SV40 chromosomes or ori-containing plasmids continued in vitro, whereas DNA synthesis in the bulk of SV40 replicative intermediate DNA (RI) that had initiated replication in vivo was rapidly inhibited. This resulted in accumulation of early RI in which most nascent DNA was localized within a 600- to 700-base-pair region centered at ori. Accumulation of early RI was observed only under conditions that permitted initiation of SV40 ori-dependent, T-antigen-dependent DNA replication and only when aphidicolin was added to the in vitro system. Increasing aphidicolin concentrations revealed that DNA synthesis in the ori region was not completely resistant to aphidicolin but simply less sensitive than DNA synthesis at forks that were farther away. Since DNA synthesized in the presence of aphidicolin was concentrated in the 300 base pairs on the early gene side of ori, we conclude that the initial direction of DNA synthesis was the same as that of early mRNA synthesis, consistent with the model proposed by Hay and DePamphilis (Cell 28:767-779, 1982). The data were also consistent with initiation of the first DNA chains in ori by CV-1 cell DNA primase-DNA polymerase alpha. Synthesis of pppA/G(pN)6-8(pdN)21-23 chains on a single-stranded DNA template by a purified preparation of this enzyme was completely resistant to aphidicolin, and further incorporation of deoxynucleotide monophosphates was inhibited. Therefore, in the presence of aphidicolin, this enzyme could initiate RNA-primed DNA synthesis at ori first in the early gene direction and then in the late gene direction, but could not continue DNA synthesis for an extended distance.
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PMID:Initiation of simian virus 40 DNA replication in vitro: aphidicolin causes accumulation of early-replicating intermediates and allows determination of the initial direction of DNA synthesis. 302 13

The Autographa californica nuclear polyhedrosis virus (AcMNPV) DNA polymerase gene was identified with the aid of an oligonucleotide probe corresponding to an amino acid sequence conserved among viral DNA polymerases of other virus families. A 3.6-kb pair region of the AcMNPV DNA, from 39.5 to 42.5 map units (m.u.), was sequenced and an open reading frame (ORF) of 2994 bp was observed. From the first ATG of this ORF, a translation product of 984 amino acids (Mr 114,310) was predicted. Amino acid sequence similarities to other viral DNA polymerases were found. Transcription was analyzed by Northern RNA blot analysis and nuclease protection studies of RNA:DNA hybrids. The ORF is transcribed in the counterclockwise direction as a 3-kb RNA. Transcripts appear to initiate at two differently regulated sites (ca. -120 and -212 bp) upstream of the initiating ATG (+1,+2,+3) and to be polyadenylated at a single site near a signal (A2UA3) which overlaps the translational termination signal (UAA) at +2952. Transcripts were observed only during a narrow window between 2 to 8 hr postinfection (p.i.) with maximum expression between 4 and 6 hr p.i. Polymerase gene transcripts were observed in the presence of the protein synthesis inhibitor cycloheximide which also blocked the shut-off of these early transcripts. Aphidicolin, an inhibitor of both viral and host DNA polymerases, inhibited polymerase gene transcription suggesting a unique regulation involving DNA replication.
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PMID:The location, sequence, transcription, and regulation of a baculovirus DNA polymerase gene. 305 78

Aphidicolin-resistant mutants (Aphr) of Bacillus subtilis bacteriophage phi 29 were isolated after mutagenesis with hydroxylamine. Efficiency of plating (e.o.p.) of the resistant mutants was not reduced at 500 microM aphidicolin, although e.o.p. of wild type phi 29 was less than 10(-5) at the same concentration of aphidicolin. By recombination and complementation analyses, both sites of the mutations, aph-71 and aph-101, of Aphr71 and Aphr101, respectively, were mapped in gene 2 which encodes phi 29 DNA polymerase. The activity of wild type phi 29 DNA polymerase, in a partially purified fraction, was inhibited by aphidicolin. DNA polymerases from Aphr71 and Aphr101, prepared in the same manner as that of wild type, were resistant to the drug. These results indicate that the acquisition of the aphidicolin resistance of Aphr71 and Aphr101 of bacteriophage phi 29 results from a structural alteration of phi 29 DNA polymerase which reduces sensitivity to aphidicolin.
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PMID:Aphidicolin-resistant mutants of bacteriophage phi 29: genetic evidence for altered DNA polymerase. 308 58

Aphidicolin, an inhibitor of DNA polymerase alpha, blocks DNA synthesis and cell division in sea urchin embryos. The effects of this inhibition appear to be stage dependent. Blastulae treated with aphidicolin before the thickening of the vegetal plate undergo developmental arrest prior to gastrulation. The extent of inhibition of DNA synthesis varies from 60 to 93% in these embryos. However, when aphidicolin is added after the vegetal plate has thickened, development continues normally through pluteus formation, even though DNA synthesis is inhibited by greater than or equal to 90% and cell division has ceased. These observations indicate that, from the vegetal plate stage onward, morphogenesis and overt differentiation are independent of DNA synthesis and cell division.
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PMID:The effects of aphidicolin on morphogenesis and differentiation in the sea urchin embryo. 309 64

Treatment of human neoplastic cells with dacarbazine both inhibits DNA synthesis and induces damage in the DNA. Lysis of cells in dilute alkali and subsequent electrophoretic analysis of the isolated DNA show that the DNA of treated cells includes a high molecular weight component and a population of 2-10-kilobase single-stranded DNA fragments while untreated cells contain only high molecular weight DNA. When DNA is pulse-labeled at the beginning of the dacarbazine treatment high amounts of small DNA fragments are seen but no labeled high molecular weight DNA. Moreover the DNA fragments are not formed in cells which are treated with aphidicolin before the addition of dacarbazine. Aphidicolin is a specific inhibitor of DNA polymerase alpha, the enzyme responsible for the replicative synthesis of DNA. We conclude that dacarbazine damages DNA only in cells which are synthesizing new DNA.
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PMID:Prevention of dacarbazine damage of human neoplastic cell DNA by aphidicolin. 309 6

Aphidicolin, a specific and reversible inhibitor of DNA polymerase alpha, was examined as a potential tool to evaluate the relationship between proliferative and differentiative events in Friend erythroleukemia cell (FELC) maturation. Since FELC can be induced to differentiate along the erythrocytic pathway with a variety of inducing agents, the effects of aphidicolin were tested on proliferating FELC and cells which were induced to differentiate with the potent inducer, hexamethylene bisacetamide (HMBA). Exposure of FELC to aphidicolin resulted in unbalanced growth within 24 h, as reflected by abnormally large cells, compared with untreated cells. In the presence of 10 or 50 microM aphidicolin, 75-90% of cells became differentiated (benzidine+ cells) within 48 h, although by 72 h cells treated with aphidicolin were non-viable as determined by trypan blue staining. A wider range of aphidicolin concentrations was tested in an effort to determine the optimal concentration of aphidicolin that maximally induced differentiation with minimal loss of cell viability. Continuous exposure of FELC from 24-96 h with doses of aphidicolin ranging from 0.5 to 50 microM was more effective for differentiation induction than was short-term exposure (1, 2, 4, 12 h) to the drug, although 1 h of exposure significantly (p less than 0.01) increased differentiation (28.1 +/- 7.8%) compared with untreated cells (2.7 +/- 1.0%). When cells were treated with HMBA (5 mM) and aphidicolin (1, 5, 10 microM), in combination, aphidicolin shifted the time of onset of differentiation from 72 to 48 h, but did not act synergistically or additively with HMBA; nor was the induction effect of aphidicolin changed by HMBA. In contrast, suboptimal doses of aphidicolin (0.5 microM) in combination with HMBA (2.5 mM) produced an additive effect on FELC differentiation. In addition, [3H]thymidine experiments demonstrated that aphidicolin reversibly blocked FELC in S phase and at G1-S interface of the cell cycle. These results indicate that aphidicolin can induce the differentiation of FELC, and that a complete round of replicative DNA synthesis is not required for differentiation to occur.
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PMID:Effect of aphidicolin on Friend erythroleukemia cell maturation. 310 67


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