Cloned fragments of human adenovirus type-12 DNA.

The following restriction endonuclease fragments of human adenovirus type 12 (Ad12) DNA have been cloned in plasmid or bacteriophage lambda vectors using standard protocols: the EcoRI-A*, -B, -D, -E, and -F fragments, the BamHI-B, -C, -D, -F, -G, -H, and -I fragments, the HindIII-F and -I fragments, and the PstI-A, -D, -F, -G, and -H fragments. The EcoRI-A* fragment comprises the right terminal 5 kb of Ad12 DNA including the terminal 143 bp.

Expression of viral DNA in adenovirus type 12-transformed cells, in tumor cells, and in revertants.

The expression as cytoplasmic RNA of integrated human adenovirus type 12 (Ad12) DNA in transformed and tumor cell lines and in revertants was investigated. The transformed and tumor cells contained multiple copies of the viral genome, 3 to 22 copies per cell in different cell lines. The integrated Ad12 DNA molecules persisted intact or nearly intact and in most cases colinear with the virion DNA.

Methylation of adenovirus genes in transformed cells and in vitro: influence on the regulation of gene expression?

An inverse correlation has been described between the levels of DNA methylation in specific segments of adenovirus DNA integrated into the genomes of transformed and tumor cells and the extent to which these segments are expressed as messenger RNA. In the adenovirus type 2 (Ad2)-transformed hamster cell lines HE2 and HE3, the virus-specific DNA binding protein (DBP) is not expressed, and the DNA in the DBP gene is completely methylated in all 5'-CCGG-3' sites. At least part of the late promoter/leader sequence of the DBP gene is present in cell lines HE2 and HE3.

An unusual symmetric recombinant between adenovirus type 12 DNA and human cell DNA.

On purification of human adenovirus type 12 (Ad12) by equilibrium sedimentation in CsCl density gradients, two bands of particles, Ad12-3 and Ad12-3a, are observed. The particles from band Ad12-3a contain a recombinant of human host cell DNA and of Ad12 DNA. The human cell DNA sequences contain repetitive DNA recurring 200 to 500 times in cellular DNA.

Integration of adenovirus type 2 DNA at a limited number of cellular sites in productively infected cells.

Several experimental findings supported the notion that adenovirus type 2 DNA is covalently linked to cellular DNA in productively infected human cells. Although the significance of this linkage for the process of viral replication is unknown, the recombination of viral DNA with host DNA may simply reflect the efficacy of the recombination systems in mammalian cells. In this report, evidence is presented for the existence of selective sites of recombination between viral and host DNAs.

Surface charge patterns as recognition signals in nucleic acid molecules.

Surface charge patterns of nucleic acid or protein molecules are caused by weak charges and are ultimately dependent upon the nucleotide or amino acid sequence, respectively. It is proposed that these charge patterns play a decisive role in nucleic acid-protein interactions. Although charge patterns are sequence related, they cannot be read off the sequence directly as yet.

Revertants of adenovirus type 12-transformed hamster cell line T637 as tools in the analysis of integration patterns.

Spontaneously arising morphological revertants of the adenovirus type 12 (Ad12)-transformed hamster cell line T637 had been previously isolated, and it had been demonstrated that in these revertants varying amounts of the integrated Ad12 genome were eliminated from the host genome. In this report, the patterns of persistence of the viral genome in the revertants were analyzed in detail. In some of the revertant cell lines, F10, TR3, and TR7, all copies of Ad12 DNA integrated in line T637 were lost.

Integration sites of adenovirus type 12 DNA in transformed hamster cells and hamster tumor cells.

The patterns and sites of integration of adenovirus type 12 (Ad12) DNA were determined in three lines of Ad12-transformed hamster cells and in two lines of Ad12-induced hamster tumor cells. The results of a detailed analysis can be summarized as follows. (i) All cell lines investigated contained multiple copies (3 to 22 genome equivalents per cell in different lines) of the entire Ad12 genome.

DNA methylation and viral gene expression in adenovirus-transformed and -infected cells.

The level of DNA methylation in adenovirus type 2 (Ad2) and type 12 (Ad12) DNA was determined by comparing the cleavage patterns generated by the isoschizomeric restriction enzymes HpaII and MspI. As previously reported virion DNA of Ad2 and Ad12 is not methylated. Parental or newly synthesized Ad2 DNA in productively infected human KB or HEK cells is not methylated either, nor is the integrated form of Ad2 DNA in productively infected cells.

Infectious DNA from Autographa californica nuclear polyhedrosis virus.

Cells from the lepidopteran Spodoptera frugiperda can be successfully transfected in culture with the DNA from Autographa californica nuclear polyhedrosis virus (AcNPV). The calcium chloride precipitation procedure has been used in conjunction with dimethyl sulfoxide treatment of the transfected cells. The highest specific infectivity observed was 6.

Adenovirus type 12-induced rat tumor cells of neuroepithelial origin: persistence and expression of the viral genome.

Four cell lines derived from adenovirus type 12-induced rat brain tumors were studied. The polyploid cells displayed neuroepithelial characteristics and were transplantable into syngeneic rats and nude mice. In tissue culture the cells grew in monolayers and multilayers.

Methylation of integrated viral DNA sequences in hamster cells transformed by adenovirus 12.

Data have been presented which demonstrate that the Ad12 DNA sequences integrated into the DNA of four lines of Ad12-transformed hamster cells are extensively methylated. Methylation in mammalian-cell DNA presumably occurs at 5'-CCGG-3' sequences mainly at the internal C residue. The majority, though not all, of the 5'-CCGG-3' sequences present in integrated Ad12 DNA are methylated.

Viruses as tools for studies on the molecular biology of mammalian cells.

1. Multiple copies of intact adenovirus type 12 (Ad12) DNA are integrated into the DNA of Ad12-transformed hamster and Ad12-induced rat brain tumor cells. Free viral DNA is not present in the Ad12-transformed lines investigated.

Methylation of integrated adenovirus type 12 DNA sequences in transformed cells is inversely correlated with viral gene expression.

The adenovirus type 12 (Ad12) DNA sequences integrated into the DNA of four lines of Ad12-transformed hamster cells are extensively methylated. Methylation in mammalian cell DNA is believed to occur predominantly at 5'-C-G-3' sequences. The majority, although not all, of the 5'-C-C-G-G-3' sequences present in integrated Ad12 DNA are methylated.

Infection of Spodoptera frugiperda cells with Autographa californica nuclear polyhedrosis virus II. The viral DNA and the kinetics of its replication.

The kinetics of replication of Autographa californica nuclear polyhedrosis virus (AcNPV) DNA in Spodoptera frugiperda cells in culture were studied. Viral DNA replication started at about 5 hr postinfection, the rate of viral DNA replication reached a maximum at about 18 hr postinfection and thereafter decreased. Parental viral DNA could be detected within the first hour postinfection in the total intracellular DNA by the Southern technique.

Infection of Spodoptera frugiperda cells with Autographa californica nuclear polyhedrosis virus I. Synthesis of intracellular proteins after virus infection.

The replication of Autographa californica nuclear polyhedrosis virus (AcNPV) in Spodoptera frugiperda cells in culture has been studied with different methods. The first virus-induced polypeptides (with molecular weights of 46K, 30K, 29K) in infected cells appeared at 3 hr postinfection. Viral DNA synthesis started at about 5 hr postinfection.

Transcription of the genome of adenovirus type 12. V. Kinetic and size-class analyses of nuclear and messenger RNA in productive infection.

In human KB cells productively infected with adenovirus type 12 (Ad12) the kinetics of synthesis of Ad12-specific RNA were investigated. Using 2 h labelling periods, striking differences were observed in the time course and the rate of synthesis of virus RNA comparing the nuclear, cytoplasmic and polysomal RNA fractions. A protracted onset of virus RNA synthesis was followed by an exponential increase between 15 and 40 h p.