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Biomedical subjects

V Daniel

Publications and source records attributed to V Daniel.

At least 145 records · Page 8Linked to original sources

Release of immature cells from the thymus during solid tumor growth: identification by assay of TdT activity.

In vivo anti-tumor activity of spleen cells from C3H/eb mice bearing a syngeneic fibrosarcoma was shown previously to decline to an undetectable level and be replaced by tumor-enhancing activity as tumor growth proceeds. In the light of our findings that thymocytes in the early stages of thymic processing can bring about tumor enhancement, we postulated that premature release of thymocytes and their accumulation in the spleen might account for the loss of the anti-tumor response. In the present experiments an injection of thymocytes did in fact cancel the anti-tumor response of reactive splenocytes from tumor-bearing mice. In order to determine whether premature thymocyte release occurs naturally in the tumor-bearing animals, we assayed activity of the enzyme TdT (as a marker for thymus cells) in the spleens of these mice during progressive tumor growth. Cells with TdT activity were clearly evident in the spleens of the tumor-bearing animals, were derived from the thymus, and accumulated in parallel to the loss of anti-tumour reactivity.

Animals↗

Altered regulation of cyclic AMP-dependent protein kinase in a mouse lymphoma cell line.

The ability of cyclic AMP to inhibit growth, cause cytolysis and induce synthesis of cyclic AMP-phosphodiesterase in S49.1 mouse lymphoma cells is deficient in cells selected on the basis of their resistance to killing by 2 mM dibutyryl cyclic AMP. The properties of the cyclic AMP-dependent protein kinase (ATP:protein phosphotransferase, EC 2.7.1.37) in the cyclic AMP-sensitive (S) and cyclic AMP-resistant (R) lymphoma cells were comparatively studied. The cyclic AMP-dependent protein kinase activity or R cells cytosol exhibits an apparent Ka for activation by cyclic AMP 100-fold greater than that of the enzyme from the parental S cells. The free regulatory and catalytic subunits from both S and R kinase are thermolabile, when associated in the holoenzyme the two subunits are more stable to heat inactivation in R kinase than in S kinase. The increased heat stability of R kinase is observed however only for the enzyme in which the catalytic and cyclic AMP-binding activities are expressed at high cyclic AMP concentrations (10(-5)--10(-4) M), the activities expressed at low cyclic AMP concentrations (10(-9)--10(-6) M) being thermolabile. The regulatory subunit of S kinase can be stabilized against heat inactivation by cyclic AMP binding both at 2-10(-7) and 10(-5) M cyclic AMP concentrations. In contrast, the regulatory subunit-cyclic AMP complex from R kinase is stable to heat inactivation only when formed in the presence of high cyclic AMP concentrations (10(-5)M). The findings indicate that the transition from a cyclic AMP-sensitive to a cyclic AMP-resistant lymphoma cell phenotype is related to a structural alteration in the regulatory subunit of the cyclic AMP-dependent protein kinase which has affected the protein's affinity for cyclic AMP and its interaction with the catalytic subunit.

Animals↗

Translation in vitro of rat liver messenger RNA coding for ligandin (glutathione S-transferase B).

Poly(A)-containing rat liver mRNA isolated from animals injected with phenobarbital and uninjected controls was translated efficiently in a wheat-germ system. The synthesis of ligandin (glutathione S-transferase B; glutathione transferase; RX-gluathione R-transferase, EC 2.5.1.18) was detected by immunoprecipitation with a highly purified monospecific ligandin antibody and analysis by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The extent of incorporation of [35S]methionine into ligandin in the translation system was similar for poly(A)-containing messages from un-infected animals and those treated with phenobarbital.

Animals↗

In vitro transcription of E. coli tRNA genes.

Transcription of tRNA genes carried by transducing bacteriophages phi80psu3+ (tRNA1Tyr) and lambdah80T (tRNA2Tyr, tRNA2Glysu36+, tRNA3Thr) was studied in vitro in a system consisting of whole bacteriophage DNA and purified RNA polymerase. In contrast to unusual requirements for tRNA1Tyr gene transcription from DNA fragments, the transcription on whole bacteriophage DNA was found to be relatively not salt sensitive, did not require glycerol and rifampicin-resistant complexes with RNA polymerase were formed in the absence of nucleoside triphosphates. Termination factor rho stimulated the transcription of the tRNA genes as well as that of 4S RNA on lambdah80T DNA template. The stimulatory effect of rho was abolished by rifampicin and seems to be due to the release of RNA polymerase and reinitiation of transcription.

DNA-Directed RNA Polymerases↗

In vitro synthesis of tRNA precursors and their conversion to mature size tRNA.

Two Escherichia coli tRNA gene clusters, tRNA1Tyr (su3+ and su3-) and tRNA2Tyr, tRNA2Gly (su+36), tRNA3Thr, were transcribed in a purified in vitro system. Evidence indicates that the adjacent tRNA genes are transcribed together as a common precursor of large size, which, on incubation with crude cell extracts, yields mature tRNA molecules.

Base Sequence↗

Induction of cytolysis of cultured lymphoma cells by adenosine 3':5'-cyclic monophosphate and the isolation of resistant variants.

Cultured mouse lymphosarcoma cells are killed on exposure to 0.1 mM N(6),O(2')-dibutyryl-adenosine 3':5'-cyclic monophosphate. A population of cells resistant to the killing effect of dibutyryl cyclic AMP at concentrations as high as 1 mM was selected. The growth characteristics of the resistant cells were similar to those of the sensitive parental line. However, the resistant cells contain less cytoplasmic cyclic AMP-binding proteins and decreased cyclic AMP-stimulated protein kinase activity. It is proposed that transition from sensitivity to resistance to dibutyryl cyclic AMP in lymphoma cells is connected with a modification of the cyclic AMP-binding protein, which appears to be the regulatory subunit of the cyclic AMP-activated protein kinase.

Animals↗

Purification and in vitro transcription of a transfer RNA gene.

A gene specifying tyrosine transfer RNA has been purified and transcribed in vitro. The purification procedure made use of two specialized transducing phages carrying the tRNA(Tyr) gene of Escherichia coli inserted into their DNA in opposite orientations. The separated heavy strands of the two phages were annealed and the single-stranded tails of the resulting hybrid were removed by digestion with Neurospora endonuclease. The size of the purified double-stranded structures was determined by electron microscopy. These isolated duplexes served as template for the in vitro transcription of tRNA(Tyr)-like molecules.

Centrifugation, Density Gradient↗