The adenovirus type 12 - mouse cell system: permissivity and analysis of integration patterns of viral DNA in tumor cells.

The integration patterns of persisting adenovirus type 12 (Ad12) DNA were analyzed in two Ad12-induced tumors of Balb/c and CBA/J mice and in one tumor cell line derived from an Ad12-induced retinoblastoma of C3H origin. In all three tumors the Ad12 genome was integrated colinearly and various copy numbers of viral DNA were found. Analysis of the Ad12 integration patterns revealed relatively simple offsize band patterns regardless of Ad12 copy numbers. The degree of methylation at the 5''-CCGG-3'' sites in the inserted Ad12 genome was determined using the isoschizomeric restriction endonuclease pair HpaII and MspI. Methylation was rather incomplete in the primary tumor tissues but almost complete in the retinoblastoma line carried in culture for many passages. The levels of expression of the viral genome in the Balb/c tumor and in the retinoblastoma line were determined by in vitro translation of RNA isolated from these cells and selected with appropriate restriction endonuclease fragments of Ad12 DNA. In both instances the 59 K, 19 K, and 17 K proteins of the E1b region were expressed. Proteins of the E1a region appeared very faint in the size class between 22 K and 42 K. The permissivity of Ad12 and the replication of Ad12 DNA in mouse cells were investigated by blotting restricted DNA from cells soon after, and a long time after, infection and by hybridization with 32P-labeled Ad12 DNA. Neither primary mouse kidney cells nor the established L929 mouse cell line supported viral DNA replication. These results raise the question to what extent host cell factors determine Ad12 DNA replication in mammalian cells.     

Genetic analysis of human lymphocyte proteins by two-dimensional gel electrophoresis: 3. Frequent occurrence of genetic variants in some abundant polypeptides of PHA-stimulated peripheral blood lymphocytes

Summary The 100 or so most intensely Coomassie blue-stained polypeptides from PHA-stimulated peripheral blood lymphocytes were analyzed by two-dimensional electrophoresis in combination with family and population studies. Besides polymorphic lymphocyte cytosol 64k polypeptide reported previously, genetic variants were frequently observed in three polypeptides with molecular weights of 100,000, 49,000, and 40,000. All of them occur in the cytosol. These variant polypeptides are charge variants, because they are separated in the isoelectric focusing dimension. It is indicated by family and population studies and cell distribution analysis that the polypeptide with a molecular weight of 100,000 shows a genetic polymorphism determined by two alleles at a new autosomal locus, as described in the following paper. Family and population studies also suggest that a genetic polymorphism defined by alleles at an autosomal locus is present in each of the polypeptides with molecular weights of 49,000 and 40,000. In contrast to the previous reports of the extremely restricted genetic variability of the 100 or so most abundant fibroblast polypeptides, the present data indicate that common genetic variants are present at least in four of the 100 or so most intensely Coomassie blue-stained lymphocyte polypeptides. The result also shows that careful side-by-side comparison of two-dimensional electrophoresis patterns among both parents and their children is an effective method to detect genetic variant polypeptides.     

Dehydrodieugenol and its monomethyl ether from Virola carinata

The constituents of Virola carinata were established as dehydrodieugenol, its monomethyl ether and sitosterol.     

New alkaloids from Banisteriopsis caapi

Three new alkaloids isolated from Banisteriopsis caapi, were identified as harmic amide (1-carbamoyl-7-methoxy β-carboline), acetyl norharmine (1-acetyl-7-methoxy β-carboline) and ketotetrahydronorharmine (7-methoxy-1,2,3,4-tetrahydro-1-oxo-β-carboline)     

In vitro construction of different oligomeric forms of lambdadv DNA and studies of their transforming activities.

Plasmid lambdadv1, which is in a dimeric form, was converted to a linear monomer duplex by the action of EcoRI restriction endonuclease that incises at a unique site in this plasmid genome. The resulting products were then joined by Escherichia coli DNA ligase to produce molecules with various oligomeric forms, and from these monomeric, dimeric, or trimeric circular molecules were purified. By transformation of cells with these DNAs, clones were obtained that carried lambdadv1 in a monomeric or dimeric form. The former type of clones have not been generated in vivo, except for one in a different host strain, and carriers of timeric or tetrameric lambdadv1's have not been obtained so far. It was observed that a considerable fraction of these oligomeric circular DNAs were converted to lower oligomers (e.g., from trimer to dimer) during transformation. The characteristics of the monomeric lambdadv1 carriers obtained were compared with those of dimeric lambdadv1 carriers. The stabilities of the plasmids of the two forms were the same. However, the monomeric plasmid carriers were less tolerant to lambdavir phage infection and perpetuated about 30% less plasmid genomes in monomer units. Furthermore, dimeric plasmid carriers appeared spontaneously and accumulated in cultures of the monomeric lambdadv1 carriers.     

New organic bases from amazonian Banisteriopsis caapi

Three new bases were isolated from Banisteriopsis caapi; they are harmine N-oxide, harmic acid methyl ester (methyl 7-methoxy-β-carboline 1-carboxylate) and harmalinic acid (7-methoxy-3,4-dihydro-β-carboline 1-carboxylic acid).