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H Ariga

Publications and source records attributed to H Ariga.

At least 127 records · Page 7Linked to original sources
PubMed1988

Cloning of the p53-dependent origin of cellular DNA replication.

We have recently reported that the c-myc protein may promote cellular DNA replication by binding to the origin of DNA replication (ori) and that an origin of human DNA replication which can autonomously replicate in human cells was cloned as a binding sequence of c-myc protein (Iguchi-Ariga et al., 1987). Here we report that cellular tumor antigen p53 may also participate in cellular DNA replication and another origin of DNA replication was cloned as a possible p53-binding sequence. The sequence could autonomously replicate in Raji cells which express p53 at a high level but not in HL-60 cells in which the coding gene for p53 is largely deleted. Little homology of the sequences was found between c-myc protein-binding ori and p53-binding ori. This suggests that c-myc protein and p53 may independently recognize different ori in chromosomal DNA.

Animals↗
PubMed1988

Adenosine deaminase (ADA) overproduction associated with congenital hemolytic anemia: case report and molecular analysis.

We report the fourth case of adenosine deaminase (ADA) overproduction associated with hereditary nonspherocytic hemolytic anemia and the molecular analysis of this anomaly. The proband was a 10-year-old Japanese boy, who had an episode of erythroblastosis fetalis during the perinatal period. The red cell ADA activity showed a 110-fold increase and the red cell ATP level was about 64% of the comparably reticulocyte-rich controls, but the lymphocyte ADA activity was within the normal range. Western blotting of partially purified ADA from red cells revealed an increased amount of enzyme in the patient's red cells. No gene amplification or gene rearrangement was found by Southern blot analysis, and no increase of ADA mRNA in reticulocyte RNA was detected by dot blot analysis using ADA cDNA. We constructed a genomic DNA library and obtained three clones containing the 5'-promoter region of ADA gene. The 2.2 kb ADA promoter DNA fragment of these clones was fused to the chloramphenicol acetyl transferase (CAT) gene, and transfected to human erythroid cell line K562, and assayed for CAT activity. One of the clones, pADOP 2 cat, expressed about 2.6 times higher CAT activity than the normal ADA promoter fused to CAT gene in K562, but such enhancement was not seen in human non-erythroid cell lines; HL 60 and Raji. From these results, it is most likely, though not conclusive, that the 5' promotor fragment of the ADA gene of the patient was responsible for the cell-specific enhancement of protein synthesis.

Adenosine Deaminase↗
PubMed1987

Dissociation between the expression of IL 2 receptor and IL 2 receptor mRNA in the antigen-specific T cell clone stimulated by the specific antigen with B cell APC.

An antigen-presenting capacity of B cells was analyzed in comparison with that of whole spleen cells in terms of IL 2R expression of a human IgG-specific T cell clone, 24-2C.3. When the clone was stimulated with the antigen and irradiated spleen cells, the clone expressed an adequate amount of IL 2R on the surface in flow cytometric analysis. On the other hand, the clone expressed the receptor only poorly, being stimulated with B cell APC. The findings were confirmed by Western blot analysis on the clone solubilized by Nonidet P-40 and also by fluorescent antibody technique on the fixed clone. 24-2C.3 clone stimulated with B cell APC was also suggested not to secrete IL 2R in culture medium. In dot blot hybridization analysis performed with cDNA probe, however, 24-2C.3 clone stimulated with B cell APC was shown to express a comparable amount of IL 2R mRNA to the clone stimulated with whole spleen cells as APC. The mRNA expressed in the clones stimulated with B cells and with spleen cells could not be distinguished from each other in Northern blot analysis. The culture supernatant of 24-2C.3 clone incubated with the antigen and irradiated spleen cells was shown to enhance the expression of IL 2R on the clone stimulated with B cell APC. These results suggest that there is some defect in the antigen-presentation capacity of B cells in terms of the elicitation of the intracellular signal for IL 2R expression in T cells and that the humoral factor(s) produced in the presence of splenic adherent cells could supply the defect.

Animals↗
PubMed1987

c-myc protein can be substituted for SV40 T antigen in SV40 DNA replication.

Replicating activity of SV40 origin-containing plasmid was tested in human cells as well as in monkey CosI cells. All the plasmids possessing SV40 ori sequences could replicate, even in the absence of SV40 T antigen, in human HL-60 and Raji cells which are expressing c-myc gene at high level. The copy numbers of the replicated plasmids in these human cells were 1/100 as high as in monkey CosI cells which express SV40 T antigen constitutively. Exactly the same plasmids as the transfected original ones were recovered from the Hirt supernatant of the transfected HL-60 cells. Furthermore, replication of the SV40 ori-containing plasmids in HL-60 cells was inhibited by anti-c-myc antibody co-transfected into the cells. These results indicate that the c-myc protein can be substituted for SV40 T antigen in SV40 DNA replication.

Animals↗
PubMed1987

Possible function of the c-myc product: promotion of cellular DNA replication.

We have recently cloned a plasmid, pARS65, containing the sequences derived from mouse liver DNA which can autonomously replicate in mouse and human cells (Ariga et al., 1987). In this report, we show that replication of pARS65 in HL-60 cells can be inhibited by co-transfection with anti-c-myc antibody. In an in-vitro replication system using HL-60 nuclear extract, pARS65 functioned as a template. This in-vitro replication was also blocked by addition of anti-c-myc antibody. Specific binding activity of the c-myc product to pARS65 was detected by an immunobinding assay, suggesting that the c-myc protein promotes DNA replication through binding to the initiation site of replication. This has been substantiated using the antibody to help isolate a human DNA segment that can autonomously replicate in the cells.

Animals↗
PubMed1987

A simple and efficient liposome method for transfection of DNA into mammalian cells grown in suspension.

For a highly efficient plasmid transfection into mammalian cells grown in suspension, DNA was entrapped in liposomes prepared by the phosphatidylserine calcium-induced fusion method. Employing this technique, a transfection efficiency of about 2% was achieved, with 22 tk+-transformants obtained from 10(3) of mouse mammary carcinoma FM3Atk- cells transfected with a plasmid carrying the thymidine kinase (tk+) gene of the Herpes simplex virus. As compared with a previous report [Ayusawa et al., J. Biol. Chem. 258 (1983) 48-53], this transfection method was more than four orders of magnitude higher than the calcium phosphate method used for FM3Atk- cells. It was shown that the tk+ gene was integrated into the chromosomal DNA and was expressed in all the tk+-transformant clones tested. The described method could be applied to various types of DNA and cells, including those grown as monolayers.

Animals↗
PubMed1987

Autonomous replicating sequences from mouse cells which can replicate in mouse cells in vivo and in vitro.

We have already reported that the cloned mouse DNA fragment (pMU65) could replicate in a simian virus 40 T antigen-dependent system in vivo and in vitro (H. Ariga, Z. Tsuchihashi, M. Naruto, and M. Yamada, Mol. Cell. Biol. 5:563-568, 1985). The plasmid p65-tk, containing the thymidine kinase (tk) gene of herpes simplex virus and the BglII-EcoRI region of pMU65 homologous to the simian virus 40 origin of DNA replication, was constructed. The p65-tk persisted episomally in tk+ transformants after the transfection of p65-tk into mouse FM3Atk- cells. The copy numbers of p65-tk in FM3Atk+ cells were 100 to 200 copies per cell. Furthermore, the p65-tk replicated semiconservatively, and the initiation of DNA replication started from the mouse DNA sequences when the replicating activity of p65-tk was tested in the in vitro DNA replication system developed from the FM3A cells. These results show that a 2.5-kilobase fragment of mouse DNA contains the autonomously replicating sequences.

Animals↗
PubMed1986

Simian virus 40 DNA replication in vitro: purification and characterization of replication factors from mouse cells.

We have previously developed simian virus 40 (SV40) DNA replication system in vitro (Ariga and Sugano, J. Virol. 48, 481, 1983). This system is composed of human HeLa or mouse FM3A nuclear extract and cytoplasmic extract of SV40 infected CosI cells. Here FM3A nuclear extract was fractionated by DEAE Sephacel and single-stranded DNA cellulose chromatography into three components required for accurate in vitro SV40 DNA replication. One fraction (A fraction) contained DNA polymerase-primase, and the second component (B fraction) contained DNA topoisomerase. Third component was further purified to near homogenuity using DEAE-Sephacel, single-stranded DNA cellulose, and glycerol gradient centrifugation. The purified protein (named factor I) bound to the origin containing fragment of SV40 DNA. The factor I enhanced the initiation of SV40 DNA replication catalyzed by SV40 infected CosI cytoplasm alone. When all four fractions consisting of A, B fractions, factor I, and SV40 infected CosI cytoplasm were mixed together, the system was reconstituted, meaning that initiation and subsequent elongation were completed to generate the full sized daughter molecules.

Animals↗
PubMed1986

Inhibition of SV40 DNA replication in vitro by 1-N-acyl-3"-N-(trifluoroacetyl) kanamycin.

A new antiviral aminoglycoside antibiotic, 1-N-eicosanoyl-3''-N-(trifluoroacetyl) kanamycin, was found to inhibit SV40 DNA replication in vitro. Several other aminoglycoside antibiotics examined did not exhibit a significant inhibition of SV40 DNA replication. The elongation of the SV40 DNA strand was profoundly affected by this agent. The degree of inhibition was decreased by increasing the amount of DNA template, but not by increasing the amount of enzyme. The inhibition of SV40 DNA replication is attributed to the interaction between the agent and the DNA template.

Amikacin↗
PubMed1986

Autonomous replicating sequences from intron of human ras gene in a simian virus 40 T antigen dependent system.

Of the several DNA fragments present in the human lung cancer gene, 1.1 and 2.0 kilobase (kb) fragments corresponding to the intron of this gene were hybridized to a half part of the 27 nucleotides perfect palindrome present in the initiation part of replication in simian virus 40 (SV40) DNA. These two fragments cloned in pBR322 had good template activity, and the initiation of DNA replication started from the region of these fragments in an in vitro system, in which the initiation of DNA replication occurs on cloned DNA containing SV40 origin of DNA replication as described previously. Furthermore, these two clones could replicate autonomously in nuclei of SV40 transformed Cos cells, producing SV40 T antigen constitutively when the clones were transfected into Cos cells. These results show that functional SV40 origin-like sequences are present in human genomes, and they can replicate autonomously within the cells which are producing SV40 T antigen.

Antigens, Polyomavirus Transforming↗
PubMed1986

The adenovirus type 12 early-region 1B 58,000-Mr gene product is required for viral DNA synthesis and for initiation of cell transformation.

An E1B 58K mutant of adenovirus type 12 (Ad12), dl207, was constructed by the deletion of 852 base pairs in the E1B 58K coding region. The mutant could grow efficiently in 293E1 cells but not in HeLa, KB, or human embryo kidney (HEK) cells. Viral DNA replication of dl207 was not detected in HeLa and KB cells and was seldom detected in HEK cells. Analysis of viral DNA synthesis in vitro showed that the Ad12-DNA-protein complex replicated by using the nuclear extract from Ad12 wild-type (WT)-infected HeLa cells but not by using the nuclear extract from dl207-infected cells. In dl207-infected HeLa and KB cells, early mRNAs were detected, but late mRNAs were not detected. The mutant induced fewer transformed foci than the WT in rat 3Y1 cells. Cells transformed by dl207 could grow efficiently in fluid medium, form colonies in soft agar culture, and induce tumors in rats transplanted with the transformed cells at the same efficiency as WT-transformed cells. Tumors were induced in hamsters injected with WT virions but were not induced in hamsters injected with dl207 virions. The results indicate that the E1B 58K protein is required both for viral DNA replication in productive infection and for initiation of cell transformation, but not for maintenance of the transformed phenotype.

Adenovirus Early Proteins↗
PubMed1986

New class of polyomavirus mutant that can persist as free copies in F9 embryonal carcinoma cells.

A small circular DNA was found extrachromosomally in a clone of F9 embryonal carcinoma (EC) cells at high copy numbers per cell. The DNA was cloned in plasmid pUC19. Restriction endonuclease analyses of the DNA indicated that the DNA (fPyF9) was a mutant of polyomavirus (Py) DNA and had a mutation in a noncoding regulatory region. There have been many reports on the isolation of Py mutants capable of replication in undifferentiated cells. However, fPyF9 was different from other Py mutants in the following aspects: it was harbored stably as a free copy at 1 X 10(4) to 5 X 10(4) copies per cell in EC cells; it replicated in undifferentiated cells better than in differentiated cells; it was extremely rearranged in the sequences of the enhancer B domain; and it carried in the enhancer B domain three copies of an exogenous sequence which does not exist in Py strain A2. From these observations, we propose a new class of Py EC mutant which has an autonomous state similar to that of plasmid and small circular DNA in host cells.

Animals↗
PubMed1985

Cloned mouse DNA fragments can replicate in a simian virus 40 T antigen-dependent system in vivo and in vitro.

Mouse liver DNA was cut out with BamHI and cloned into YIp5, which contained the URA3 gene of Saccharomyces cerevisiae in pBR322. Of the several plasmids isolated, two plasmids, pMU65 and pMU111, could transform S. cerevisiae from the URA- to the URA+ phenotype and could replicate autonomously within the transformant, indicating that mouse DNA fragments present in pMU65 or pMU111 contain autonomously replicating sequences (ARS) for replication in S. cerevisiae. Furthermore, to determine the correlation between ARS function in yeast cells and that in much higher organisms, we tried to challenge these plasmids with the simian virus 40 (SV40) DNA replication system. Of the two plasmids tested, the EcoRI-BglII region of pMU65 could be hybridized with a chemically synthesized 13-nucleotide fragment corresponding to the origin region of SV40 DNA. Both pMU65 (the EcoRI-BglII region cloned in pBR322) and its subclone pMU65EB could replicate semiconservatively, and initiation of DNA replication started from the EcoRI-BglII region when the replicating activity of these plasmids was tested in the in vitro SV40 DNA replication system we have established before. Furthermore, pMU65 and pMU65EB could replicate autonomously within monkey Cos cells which produce SV40 T antigen constitutively. These results show that a 2.5-kilobase fragment of the EcoRI-BglII region in pMU65 contains the ARS needed for replication in the SV40 DNA replication system.

Animals↗