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Biomedical subjects

S K Arya

Publications and source records attributed to S K Arya.

At least 55 records · Page 3Linked to original sources
PubMed1984

Organization of polyoma virus DNA in mouse tumor cell lines.

Restriction mapping of polyoma virus DNA in mouse tumor cell lines gave patterns that varied with the cell line examined. These reflected differences in both the organization and the state of integration of virus genomes in the host chromosomes. (The cell lines were derived from tumors induced by polyoma virus in vivo and were propagated continuously in culture.) Two of the cell lines contained multiple copies of tandemly integrated virus genomes as well as free virus DNA molecules. Two other cell lines appeared to contain only integrated virus genomes arranged as tandem repeats. Based on restriction analysis with eleven different endonucleases, the virus DNA in one of the cell lines containing both free and integrated virus genomes was not detectably defective or hypermethylated. This is in contrast to most previously described polyoma virus transformed mouse cells. These virus genomes may, however, contain point mutations or unobserved rearrangements. The second cell line possessing free virus DNA molecules contained both nondefective and defective virus genomes. Most, if not all, defective virus genomes in this line were integrated. The two other cell lines possessing only detectable integrated virus DNA apparently contained only defective virus genomes. The defect in both cases was a small deletion (less than or equal to 0.2 kb) encompassing 0.12 map units on the physical map of polyoma virus DNA, a region coding for the proximal part of the large T antigen. Moreover, in contrast to the cell lines with free and detectably nondefective virus DNA, the virus DNA was extensively methylated in cell lines containing only integrated and defective virus genomes.

Animals↗
PubMed1984

Cyclosporin A inhibits T-cell growth factor gene expression at the level of mRNA transcription.

Cyclosporin A (CsA) is a potent immunosuppressive agent, now gaining wide application in human organ transplantation. The immunosuppressive activity of CsA is at least in part due to inhibition of lymphokine production by activated T lymphocytes. Specifically, inhibition of T-cell growth factor (TCGF; also designated interleukin 2) production appears to be an important pathway by which CsA impairs T-cell function. To define further both the specificity of CsA and the level at which it interferes with lymphokine gene expression, we have studied its effects on TCGF mRNA accumulation as well as TCGF gene transcription. These studies were performed with a cloned human leukemic T-cell line (Jurkat, subclone 32), which can be induced with phytohemagglutinin and phorbol 12-myristate 13-acetate to produce large amounts of TCGF. In these cells, high levels of TCGF mRNA were present in induced but not in uninduced Jurkat cells as judged by hybridization to a cloned human TCGF cDNA probe. CsA completely inhibited induced TCGF mRNA accumulation at concentrations of 0.3-1.0 microgram/ml, whereas low levels of appropriately sized TCGF mRNA were present at 0.01 microgram/ml. In nuclear transcription experiments, CsA inhibited the synthesis of TCGF transcripts in a dose-dependent manner with complete inhibition at a concentration of 1 microgram/ml. In contrast, CsA did not inhibit the expression of two other inducible genes, TCGF receptor and HT-3. Further, HLA gene expression was also less affected than TCGF in CsA-treated cells. These data suggest a relatively selective action of CsA on TCGF gene transcription.

Cell Line↗
PubMed1984

Human T-cell growth factor: partial amino acid sequence, cDNA cloning, and organization and expression in normal and leukemic cells.

The partial amino acid sequences of human T-cell growth factors (TCGFs) isolated from normal peripheral blood lymphocytes and from a leukemia T-cell line (Jurkat) show that the amino-terminal sequences of the two proteins (15 residues) are identical. Oligonucleotides based on the published Jurkat TCGF DNA sequence were used to isolate six cDNA clones of TCGF mRNA from normal lymphocytes. The predicted amino acid sequence of normal lymphocyte TCGF was identical to the sequence of the Jurkat protein, showing that the differences in biochemical properties of the two proteins result from post-translational events. Amino acid and nucleotide sequence data suggest that TCGF is derived from a precursor polypeptide that is cleaved at the amino terminus but not at the carboxyl terminus. Hybridization of the cloned lymphocyte TCGF cDNA to cellular DNA and RNA strongly suggested that the TCGF gene is expressed as a single mRNA species from a single-copy gene. No differences in the organization of the TCGF gene in normal, leukemic, and human T-cell leukemia/lymphoma virus-infected cells was detected regardless of whether they produce TCGF or not.

Amino Acid Sequence↗
PubMed1984

Transcriptional regulation of a tumor promoter and mitogen-inducible gene in human lymphocytes.

Tumor-promoting phorbol esters affect a variety of cellular functions which may underlie tumor promotion. We isolated from human lymphocytes a cDNA clone whose gene is inducible by the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate as well as by the T-cell mitogen phytohemagglutinin. Nuclear transcription experiments suggested that this induction is primarily caused by the increased transcription of the gene. It is interesting that this gene is expressed constitutively in human T-cell leukemia virus-infected mature T cells. The results support the notion that 12-O-tetradecanoyl-phorbol-13-acetate can affect cellular functions by causing transcriptional activation of specific genes.

Cells, Cultured↗
PubMed1984

Molecular characterization of human T-lymphotropic leukemia virus type III associated with the acquired immunodeficiency syndrome.

A T-lymphotropic retrovirus with cytopathic but not immortalizing activity has been isolated repeatedly from patients with acquired immune deficiency (AIDS) or lymphadenopathy syndrome (LAS) and successfully transmitted to a T-cell line (HT) for continuous production. Seroepidemiology data and the OKT4 tropism and cytopathogenicity of this virus indicate it is the etiological agent of AIDS. We have cloned HTLV-III genomes using three approaches: (1) cDNA clones were obtained from a cDNA plasmid library constructed from RNA of purified virions using oligo (dT) primers; (2) unintegrated provirus clones were obtained from Hirt supernatants of acutely infected H9 cells using virus from H9/HTLV-III; (3) clones of integrated provirus with flanking cellular sequences were obtained from a genomic DNA library of H9/HTLV-III. Analyses of these clones show that the HTLV-III genome is similar in size to those of HTLV-I and HTLV-II and contains a gene that functions as a transcriptional activator. Different isolates of HTLV-III display greater polymorphism than different isolates of HTLV-I among each other, possibly due to the highly replicative nature of HTLV-III. Viral sequences could be detected in fresh lymph node tissues of some AIDS patients, but even in the positive samples the number of infected cells is small (less than 1%). In both fresh tissues that are positive for viral sequences and HTLV-III infected cell lines, a substantial amount of unintegrated viral DNA is present in addition to integrated provirus. This is an unusual finding for retroviruses but may be significant in the cytopathicity of HTLV-III as has been proposed for some avain retroviruses.

Acquired Immunodeficiency Syndrome↗
PubMed1984

Dexamethasone-mediated inhibition of human T cell growth factor and gamma-interferon messenger RNA.

Glucocorticoids suppress the proliferation of human T lymphocytes. Activated T lymphocytes require T cell growth factor (TCGF) for proliferation. TCGF is produced by a subset of T lymphocytes, and this production is regulated at the TCGF mRNA level. Dexamethasone, a synthetic glucocorticoid, strongly inhibits the synthesis of TCGF mRNA in human normal peripheral blood lymphocytes stimulated in culture with phytohemagglutinin. It also inhibits the accumulation of gamma-interferon mRNA in these cells. This dual effect may in part explain some of the immunosuppressive and anti-inflammatory effects of glucocorticoids.

Cell Line↗
PubMed1982

Differential expression of the amv gene in human hematopoietic cells.

Total cellular RNAs from a variety of fresh and culture-derived human hematopoietic neoplastic cell types at various stages of differentiation and human sarcoma, carcinoma, melanoma, and glioblastoma cell lines were enriched for poly(A)- containing sequences, fractionated by gel electrophoresis, and blot hybridized to a cloned DNA probe containing the transforming sequences (v-amv) of avian myeloblastosis virus (AMV), a virus known to cause myeloid leukemias in chickens. Expression of RNA sequences homologous to AMV was detected in all immature myeloid and lymphoid T cells in addition to the single erythroid cell line examined, but not in mature T cells or in B cells, including lymphoblast cell lines derived from patients with Burkitt lymphoma. In addition, induction of the cell line HL60, a promyelocytic leukemia line, to differentiate with dimethyl sulfoxide or retinoic acid resulted in a reduction of the level of expression of the human cellular gene c-amv homologous to v-amv. There was no detectable c-amv mRNA in any of the solid tumor cell lines examined. Thus, expression of the human c-amv gene could be correlated with the stage of differentiation of different hematopoietic cell types determined by morphologic and marker studies. Expression of c-amv could not be correlated with the extent of methylation in HL60 and in HL60 induced to differentiate with dimethyl sulfoxide.

Carcinoma↗
PubMed1982

Mouse mammary tumour virus and polyoma virus information in mammary tumours of athymic mice inoculated with polyoma virus.

Polyoma virus inoculation of athymic mice results in the development of mammary tumours with a much higher incidence than the development of salivary gland tumours, the latter being the most common for immunocompetent normal mice. The possibility existed that polyoma virus might act as a co-carcinogen in activating the expression of mouse mammary tumour virus (MMTV). Molecular hybridization studies, however, showed that the mammary tumour development was accompanied by neither the amplification of MMTV genomic sequences nor by their more extensive transcription. In contrast, tumour tissue contained about 60 to 100 copies of polyoma virus genome equivalents per cell and some of these sequences were apparently transcribed into RNA. While these results do not rule out the transient involvement of MMTV expression in mammary tumour development, it appeared that the mammary gland cells were directly transformed by polyoma virus. Apparently, polyoma virus displayed a tropism in athymic mice that was different from that in normal mice.

Animals↗
PubMed1982

Organization of polyoma virus DNA in mammary tumours of athymic mice.

Restriction mapping of polyoma virus DNA in mammary tumours of athymic mice gave patterns that varied with the tumour examined. These reflected differences in both the organization and the state of integration of virus genomes in the host chromosomes. All tumours contained tandemly integrated full-length and defective virus genomes. Some tumours also contained unintegrated virus DNA molecules, some full-length and others defective. The deletions were localized in the virus genomic sequences coding for the distal part of the large T antigen. After the first transplantation, the organization of polyoma virus genomes in tumours remained essentially unchanged through four successive transplantations. The tumour cells that initially contained free virus DNA molecules continued to possess such molecules during serial transplantations. The virus DNA molecules in transplanted tumours lacking unintegrated virus genomes were more methylated than in tumours containing unintegrated virus genomes.

Animals↗
PubMed1981

Interferon-mediated inhibition of mouse mammary tumour virus (type B) and mouse leukaemia virus (type C) in the same culture.

The sensitivity of mouse mammary tumour virus (MMTV) and murine leukaemia virus (MuLV) to interferon was tested by investigating the effect of interferon in cultures of mouse mammary tumour cells simultaneously synthesizing the two viruses. The dose-response curves showed that the extracellular production of both viruses by these cells was equally inhibited by interferon treatment. In contrast, the intracellular steady-state concentration of virus-specific RNA of both MMTV and MuLV was not significantly affected in these cells. This was the case regardless of whether total cellular RNA or polysomal poly(A)-containing RNA was examined. These results are consistent with the suggestion that interferon-caused inhibition is related to a host-mediated effect on a post-transcriptional step in virus synthesis.

Animals↗
PubMed1981

Comparison of the sequence complexity and abundance frequency of polyadenylated messenger RNA in human breast and prostate carcinoma cells.

The sequence complexity and frequency distribution of polyadenylated [poly(A)+] messenger RNA (mRNA) in human breast and prostate carcinoma cells were compared by the kinetics of homologous and heterologous mRNA:complementary DNA hybridization. The apparent total sequence complexity of human breast cell poly(A)+ RNA was estimated to be about 47,300 kilobases (kb). It appeared to be distributed in three abundance classes comprising about 14 (32 kb), 43 (2,000 kb), and 43% (45,270 kb) of the total. The poly(A)+ mRNA of human prostate cells also appeared to be organized in three abundance classes: the highest-abundance class comprising about 15% of the total with an approximate complexity of 88 kb; an intermediate-abundance class comprising about 32% of the total with a complexity of about 2,330 kb; and a low-abundance class which was about 53% of the total and had an estimated complexity of 66,840 kb. The results of heterologous hybridization reaction suggested that about 75 to 80% of the poly(A)+ mRNA sequences present in human breast cells were also present in human prostate cells. Similarly, about 75 to 80% of the sequences in human prostate cells appeared to be present in human breast cells. The sequences held in common by these two cell types appeared to be present in roughly similar relative abundance. The hybridization of kinetically fractionated abundant complementary DNA of breast or prostate cells with homologous and heterologous poly(A)+ mRNA suggested that the abundant sequences of one cell type (e.g., breast) were present in reduced abundance in the other cell type (e.g., prostate) but only by a factor of 2 to 4. Nearly all of the abundant sequences of one cell type appeared to be present in the second cell type.

Base Sequence↗
PubMed1981

Changes in retrovirus expression in mouse mammary tumor cells in culture.

Mouse mammary tumor cells (Mm5Mt/c1) produced mouse mammary tumor virus (MMTV) (type B retrovirus) at early passages in culture. They apparently produced predominently type C retrovirus(es) at late passages in culture. The decline in MMTV production during passage was not accompanied by an apparent decrease in the synthesis or accumulation of intracellular MMTV-specific RNA. As judged by the concentration of RNA (mol/titer) x half-time (sec) of hybridization of cellular RNA with MMTV complementary DNA, the intracellular concentration MMTV-specific RNA did not change significantly during passage in culture. Since late-passage RNA, particularly polysomal poly(adenylic acid)-containing RNA, yielded lower apparent maximum hybridization of MMTV complementary DNA as compared with early-passage RNA, it was possible that the complexity or base sequence of MMTV-specific RNA in late-passage cultures was not identical with that in early-passage cultures. The data on thermal transitions of hybrids were in accord with this possibility. In contrast, the steady-state level of type C virus-specific RNA in late-passage cultures was orders of magnitude higher than that in early-passage cultures. The production of type C retrovirus(es) at later passages was apparently accompanied by increased transcription of type c retrovirus DNA.

Animals↗
PubMed1980

Synthesis in high yield of complementary DNA of retroviral RNA.

Mouse mammary tumor virus RNA was transcribed in vitro with avian myeloblastosis virus reverse transcriptase in the presence of calf thymus DNA oligomers. The yield of cDNA was markedly enchanced by increasing the concentration of the enzyme as well as primers in the reaction mixture. The average size of cDNA was approximately 200 residues, and it was not affected by the concentration of deoxynucleoside triphosphates when saturating concentration of enzyme was used. Nearly all of the viral sequences were represented in cDNA and it formed hybrids of high fidelity with viral RNA. Similar results were obtained when murine leukemia virus (AKR) RNA was transcribed. These observations will be useful for synthesizing cDNA of RNAs that are not easily available in sufficient quantities.

Avian Leukosis Virus↗
PubMed1980

Interferon induced inhibition of mouse mammary tumor virus production.

Treatment of murine cells chronically infected with mouse mammary tumor virus with interferon resulted in a marked inhibition of extracellular virus production. This inhibition occurred despite the lack of effect on the transcription or translation of viral RNA. Interferon treatment did not alter the steady-state level of intracellular viral RNA as determined by hybridization of total cellular and polysomal poly(A) containing RNA with mouse mammary tumor virus complementary DNA. Additionally, analyses by polyacrylamide gel electrophoresis showed no differences in the molecular weights of the major viral proteins in extracts from control and interferon treated cultures. These results are consistent with suggestions that the interferon inhibits production of mouse mammary tumor virus at a late stage of replication.

Animals↗
PubMed1980

Simian-sarcoma-virus-related sequences in DNAs from human prostatic tissues.

Several prostatic tissue DNAs have been examined for their content of simian-sarcoma-virus-related sequences. This was done by hybridizing fragmented prostatic DNA with simian sarcoma virus complementary DNA (cDNA) synthesized in vitro. The tissue DNAs contained limited amounts of virus-related sequences. The content ranged from about 10 to 30 pg of viral DNA per milligram of cell DNA, depending on the tissue. This corresponds to approximately one viral genome equivalent per 20-100 cell genomes. A clear correlation between viral DNA content and histopathology of the tissue - normal, hyperplastic and neoplastic - has not emerged from the tissue DNAs examined thus far. The trend appeared to be for the hyperplastic tissue to contain more virus-related sequences than normal or neoplastic tissue. The melting point of hybrids of prostatic DNA and viral cDNA was 15 degrees C lower than that of native prostatic DNA. This can be interpreted to suggest that prostatic DNAs contain a distribution of short segments of sequences that hybridize with viral cDNA.

Base Sequence↗
PubMed1980

Retrovirus-like particles released by human prostatic cells.

A cell strain has been obtained from an explant culture of a human prostatic tissue. The tissue was diagnosed as benign prostatic hyperplasia. Cultured cells displayed human male karyotype and chromosome model number of 43--46. Cultures of these cells appear to release retrovirus-like entities. The extracellular particulate material obtained from the culture medium showed the following characteristics: (1) it banded at a density of 1.12--1.18 g/ml in a sucrose density gradient; (2) it contained particles with a sedimentation rate approximating that of the known retroviruses; (3) these particles contained RNA and RNA-directed DNA polymerase; (4) the RNA-directed DNA polymerase utilized poly (Cm) as a template, and (5) the particles synthesized DNA in an endogenous DNA polymerase reaction. The DNA thus synthesized was associated with RNA, some of it with high molecular weight RNA. These observations can be interpreted to suggest that human prostatic cells produce retrovirus(es). However, we were not able to detect a 60--70S RNA species in the labeled RNA of the released particles. The isolated RNA, however, displayed a 30--35S component.

Cell Line↗