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M T Kuo

Publications and source records attributed to M T Kuo.

At least 73 records · Page 4Linked to original sources
PubMed1981

Preferential damage of active chromatin by bleomycin.

Preferential sensitivity of specific genes in chromatin to the action of bleomycin in different cell types was investigated using restriction enzyme digestion of DNA from drug-treated nuclei in combination with the Southern blotting procedure. The results show that the highly active ovalbumin gene in laying hen oviduct nuclei is more sensitive than the inactive globin gene to the digestion by bleomycin. Conversely, in red blood cells, bleomycin preferentially destroys the globin gene but not the ovalbumin gene. Therefore, bleomycin, like nucleases, can preferentially destroy DNA sequences in chromatin with open configuration, although the mode of action of this drug on DNA is drastically different from those of known nucleases.

Animals↗
PubMed1980

Differential nuclease sensitivity of the ovalbumin and beta-globin chromatin regions in erythrocytes and oviduct cells of laying hen.

We have monitored the differential nuclease sensitivity of defined regions of the chicken genome in different cells using a method which combines restriction enzyme digestion and blotting to diazobenzyloxymethyl (DBM)-paper (see Ref. 11). By using different specific probes and by scanning the bands on the autoradiograms, it is possible to compare on the same blot the digestion patterns of similar-sized fragments from different regions of the genome corresponding to "active" and reference "inactive" genes. We have demonstrated the preferential sensitivity to DNaseI and micrococcal nuclease digestion of the ovalbumin gene region in hen oviduct chromatin. The beta-globin gene region (containing both an adult and an embryonic gene) is also preferentially digested by DNaseI in hen mature erythrocyte nuclei, but at a lower rate than the ovalbumin gene region in oviduct. These observations raise the possibility that there may be several types of preferential nuclease sensitivities, all characterized by increased rates of digestion but to different levels, the highest corresponding to the very actively transcribing genes.

Animals↗
PubMed1979

DNA methylation: correlation with DNase I sensitivity of chicken ovalbumin and conalbumin chromatin.

To analyse the relationship between DNA undermethylation at some sites in the ovalbumin and conalbumin gene regions (1) and the expression of these genes in chick oviduct, digestions with HhaI, which differentiates between methylated and unmethylated HhaI restriction sites, was performed on DNA isolated from chicken erythrocyte or oviduct chromatin treated with DNase I which degrades preferentially "active" chromatin. This was followed by analysis with ovalbumin- and conalbumin-specific hybridization probes. We conclude that the residual DNA methylation found at some sites of the ovalbumin and conalbumin gene regions is derived from the fraction of cells in which the chromatin of these genes is not in an "active" form. On the other hand, the ovalbumin and conalbumin sites which are partially unmethylated in erythrocyte DNA correspond to chromatin regions which are not DNase I-senitive. We have also detected a site about 1 kb downstream from the 3' end of the conalbumin gene that is hypersensitive to DNase I in all tissues tested.

Animals↗
PubMed1979

Studies in heterochromatin DNA: accessibility of late replicating heterochromatin DNA in chromatin to micrococcal nuclease digestion.

Heterochromatin DNA in cactus mouse (Peromyscus eremicus) replicates in the late S phase of cell cycle. A method of obtaining cells which contain DNA preferentially labeled at heterochromatic areas by a pulse-labeling of late replicating DNA is described. When the nuclei of P. eremicus cells containing radioactively labeled DNA in heterochromatin were digested with micrococcal nuclease and the resultant nucleosomal DNA was separated by gel electrophoresis, it was found that the repeat length of nucleosomal DNA in the heterochromatin DNA is not different from that of the bulk of the genomic DNA. Furthermore, there was no significant difference in the accessibility to digestion by micrococcal nuclease between the late replicating heterochromatin DNA and the total DNA under our digestion conditions. Two dimensional gel electrophoresis patterns of nucleosomal DNAs isolated from micrococcal nuclease digested nuclei from P. eremicus, P. collatus, and P. crinitus cells in culture were very similar. Cytogenetic data showed that these three species are different in heterochromatin but similar in euchromatin.

Animals↗
PubMed1978

Effects of neocarzinostatin on mammalian nuclei: release of nucleosomes.

When Chinese hamster cell nuclei (Line CHO) were reacted with neocarzinostatin and its DNA was analyzed on non-denaturing agarose gel electrophoresis, a series of bands with a multiplicity of 175 base pairs was obtained. A similar result was also obtained when the DNA samples were electrophoresed under denaturing gels. Our results suggest that the linker DNA between nucleosomes is the susceptible site to the drug and that neocarzinostatin can be used to study the chromatin structure.

Antibiotics, Antineoplastic↗
PubMed1978

Studies in heterochromatin DNA: characterization of transcripts synthesized in situ from C-banded preparations.

The transcript of in situ RNA synthesis using Peromyscus eremicus cells, which had been treated with HCl followed by NaOH, proved to be rich in heterochromatin specifying sequences. Such transcript, referred to as heterochromatin RNA, can be characterized biochemically to probe the organization of heterochromatin DNA. It was found that the heterochromatin RNA of P. eremicus contained non-repetitive sequences covalently linked to repetitive sequences.

Animals↗
PubMed1977

Genome analysis of Peromyscus (Rodentia, Cricetidae) VII. Localization of satellite DNA sequences and cytoplasmic poly(A) RNA sequences of P. eremicus on metaphase chromosomes.

A satellite DNA fraction from P. eremicus, having a buoyant density of 1.705 g/ml in neutral CsCl density gradients, was isolated. In situ hybridization experiments, using 3H-RNA complementary to this DNA fraction indicated that the short (heterochromatic) arms of most of the autosomes contained this sequence. Conversely, in situ hybridization using 3H-complementary DNA (cDNA) synthesized from the cytoplasmic poly (A) RNA of P. eremicus (comprising a substantial fraction of total messenger RNA) showed that the number of silver grains in the long arms (euchromatin) was significantly higher than that in the short arms. The X chromosomes showed a distinct localization pattern of both sequences.

Animals↗
PubMed1977

Effect of bleomycin on the synthesis and function of RNA.

Bleomycin inhibits cellular RNA synthesis and the inhibition is nonspecific. The ratio of polyadenylate- [poly(A)] containing RNA to non-poly(A)-containing RNA in the drug-treated human lymphocytic cells, line Wil2, was the same as that in untreated cells. Poly(A) RNA isolated from untreated cells was used as a template for reverse transcriptase to synthesize complementary DNA, which was then used as a probe to assay the sequence diversity of poly(A)RNA's from treated and untreated cells. It was found that essentially all of the poly(A) RNA's in the untreated cells were also present in the treated cells. The effect of bleomycin on the biological activity of messenger RNA (mRNA) was tested with globin mRNA in a wheat germ embryo translation system. Although bleomycin inhibited protein synthesis at high concentrations, the inhibition was not due to a modification of mRNA. This was evidenced by the fact that no decrease in the ability of mRNA to function in the test system was found when globin mRNA was pretreated with high concentrations of bleomycin followed by removal of the drug.

Bleomycin↗
PubMed1976

Abundancy and diversity of mRNA sequences in human leukocytes.

The messenger RNA populations in two haematopoietic tissues were compared with respect to number of different sequences present and their relative abundancies. This was accomplished by hybridising complementary DNA to the cytoplasmic polyadenylated RNA from which it was transcribed and to heterologous polyadenylated RNA. The hybridisation kinetics were essentially the same in the homologous hybridisation. However, in heterologous hybridisation some differences were observed in the high abundancy messengers.

Base Sequence↗
PubMed1975

Localization of cytoplasmic-membrane-associated DNA in human chromosomes.

In situ hybridization was used to localize the sites of the repeated sequences of cytoplasmic-membrane-associated DNA (cmDNA) in human chromosomes. cmDNA was mainly present in the heterochromatic regions of chromosomes 9. Although cmDNA has buoyant densities in neutral and alkaline CsCl gradients similar to those of human satellite C DNA, which also is localized in the heterochromatic region of chromosomes 9, cmDNA was not homologous to satellite C DNA as determined by DNA-complementary RNA hybridization. These results are in agreement with our previous studies, which indicated that cmDNA is a unique species of human DNA.

Binding Sites↗
PubMedDate not supplied

Murine mdr-1, mdr-2, and mdr-3 gene expression: no coinduction with the Cyp1a-1 and Nmo-1 genes in liver by 2,3,7,8-tetrachlorodibenzo-p-dioxin.

Multidrug-resistance (MDR) genes are induced in the liver of rodents treated with a variety of foreign chemicals and hepatocarcinogens. It has been reported that 2,3,6,7-tetrachlorodibenzo-p-dioxin (TCDD) might increase hepatic MDR transcripts in the Fischer rat and the C57BL/6 (B6) inbred mouse strain having the high-affinity aromatic hydrocarbon (Ah) receptor, but not in the DBA/2 (D2) strain having the low-affinity Ah receptor. These intriguing results suggest that TCDD might activate MDR gene expression by way of an Ah receptor-mediated signal transduction pathway. We have attempted to confirm these data in four inbred mouse strains: two (B6 and BALB/c) having the high-affinity Ah receptor, and two (D2 and AKR) having the low-affinity Ah receptor. The RNase protection assay was used to distinguish between the MDR1, MDR2, and MDR3 mRNAs. TCDD treatment at high (100 micrograms/kg) and low (1 mu/kg) doses, a time course from 6 to 96 hr of TCDD treatment, progeny from the B6D2F1 x D2 backcross, and transcriptional run-on experiments were performed. The Cyp1a-1 (cytochrome P1450) and Nmo-1 [NAD(P)H:menadione oxidoreductase] genes, two members of the TCDD-inducible [Ah] battery, were used as positive controls. We were unable to detect significant coinduction of MDR1, MDR2, or MDR3 mRNA with CYP1A1 mRNA or with Cyp1a-1 or Nmo-1 transcription under any conditions. Therefore, we conclude that any effects that TCDD might have on MDR expression must be substantially different from TCDD effects on genes known to be induced via the Ah receptor.

Animals↗
PubMedDate not supplied

Isolation and characterization of putative intrinsic multidrug resistant Chinese hamster ovary cells by fluorescence activated cell sorting.

Most multidrug resistant cell lines reported in the literature were established by long-term continuous exposure of cells to stepwise increasing concentrations of antitumor drugs. However, these resistant cell lines may not be relevant to the majority of clinically resistant cells. In this study, we described the establishment of doxorubicin (Dox)-resistant Chinese hamster ovary cells by repeated flow cytometric cell sorting using the intrinsic fluorescence of Dox. In each sorting, the 15% least fluorescent cells were fractionated, grown to mass culture and sorted again. Results from a total of nine sorting cycles showed that the intracellular levels of Dox in the sorted cells were inversely proportional to the number of sorting cycles. The levels of P-glycoprotein mRNA in the sorted cells were increased, but reached a plateau of 2-3 fold after the fifth sorting cycle. The sorted cells exhibited a moderate but stable multidrug-resistant phenotype. Because the procedure involved minimal exposure of cells to the drug, the isolated cells are most likely related to naturally occurring (intrinsic) MDR cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗