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UV radiation and other carcinogenic agents induce an increase in DNA-binding activity to the early domain of the simian virus 40 (SV40) minimal origin in both SV40-permissive and SV40-nonpermissive cells. The increase is due to posttranslational modification of a preexisting protein, since it occurs in the presence of cycloheximide or anisomycin. Binding of this factor is an absolute requirement for the UV-induced SV40 DNA amplification in Co631 cells in vivo. A synthetic double-stranded oligonucleotide covering the early domain sequence totally blocked the UV-induced amplification in competition experiments. Point mutants of the sequence and unrelated oligonucleotides which could not bind the factor also did not block SV40 amplification. Inhibitors of protein synthesis caused an immediate increase of both early-domain factor activity (perhaps by prolonging mRNA half-life for the factor or for a modifying enzyme) and DNA amplification. The effects of UV and cycloheximide on SV40 amplification were superaddition.

One transcription factor, AP-1, has, through the complementation of data from different research areas, more than any other one promoted our understanding of gene regulation and transduction of extracellular signals to the nucleus. In this article, we describe the constituent components of AP-1, their cooperation in binding to DNA and in transcription in vitro and our current understanding of AP-1 dependent gene regulation.

Prior to the isolation of mammalian DNA repair genes and identification of their gene products, the comparison between the bacterial SOS response and various similar reactions in mammalian cells remains rather speculative. The increasing number of observed phenomena including enhanced DNA repair, virus induction, induced cellular differentiation, and neoplastic transformation, all following DNA damage or arrest of replication, are, however, suggestive of an SOS-like system of growth control and may form an entry into this fascinating area.

Various isolated cells of an inbred mouse strain (BALB/c) differed widely in their sensitivity to gamma irradiation: fibroblasts are five times more resistant than peripheral lymphocytes. Among lymphocytes, T cells are more resistant than B cells. Cell lines derived from the primary cells conserved their radiosensitivity. Cytofluorometric measurements show that the differential reaction of a cell to gamma irradiation can be detected already 2–3 h after the irradiation event. Radiation-sensitive cells are delayed for a longer time in S phase and G2 phase of the cell cycle than radiation-resistant cells. No difference in the capacity of the cells to perform single-strand break repair, double-strand break repair or unscheduled DNA synthesis could yet be detected.

Fos and Jun co-operatively repress the fos promoter. Removal of all putative Fos/Jun binding sites from the fos promoter neither obliterates the repression by Fos/Jun in transient cotransfection experiments in NIH3T3 cells nor the turn-off kinetics of serum-induced fos expression in stably transfected NIH3T3 cells. The dyad symmetry element (DSE) suffices to subject a promoter to this type of repression. However, one of the putative Fos/Jun binding sites (-292 to -299 and thus located immediately adjacent to the DSE), determines the very low level of basal expression.

The T3 phage enzymes S -adenosyl methionine cleaving enzyme and lysozyme and the T7 lysozyme were synthesized in a deoxyribonucleic acid (DNA)-dependent, cell-free system derived from uninfected Escherichia coli. The data presented suggest that these enzymes are encoded in that portion of the DNA which is transcribed early after infection.

RNA polymerase from Escherichia coli transcribes on T7 DNA in vitro and in vivo a polycistronic messenger RNA of 2.2 × 106 molecular weight. This mRNA contains the information for synthesis of T7-phage RNA polymerase, ligase and lysozyme as was demonstrated by enzyme synthesis in vitro, using the RNA synthesized in vitro as template. Bystudying the kinetics of synthesis of these enzymes in vitro and the ultraviolet-irradiation sensitivity of enzymesynthesis rates in vivo, the location of the genes on the DNA was estimated. Phage RNA polymerase is encoded within the first half of the mRNA molecule. The information for ligase ends at approximately 5500 nucleotides (1.7 × 106 mol wt.) and the information for lysozyme is close to the 3′-end of the mRNA. The genes for T7 RNA polymerase and ligase are transcribed by the host RNA polymerase (early region), whereas the genes for endonuclease (gene 3) and for lysozyme are read by E. coli RNA polymerase and by T7-phage RNA polymerase (early/late region). The DNA beyond the termination signal for host RNA polymerase is transcribed by T7-phage RNA polymerase in the late phase (late region). Genetic mapping of mutants in the ligase and in the lysozyme genes confirm the data obtained by enzyme synthesis in vitro.

Bacteriophage φμ-4 was purified from lysates by isoelectric precipitation and CsCl gradient centrifugation. This phage displays an Escherichia coli phage lambda-like appearance in electron microscopy with polyhedrical heads and long tails, but without terminal plates and tail fibers. Bacteriophage φμ-4 is a DNA virus. The DNA has a molecular weight 26 × 10 6 daltons, a buoyant density of 1.719 g/cm 3 , a G + C-content of 58% and appears to contain no unusual base residues. The DNA strands can be separated by alkaline CsCl gradient centrifugation or annealing to poly U,G, and subsequent CsCl gradient centrifugation. DNA of phage φμ-4 is an excellent template for in vitro transcription by both E. coli RNA-polymerase and host RNA-polymerase ( Bacillus stearothermophilus ). There are 15 polymerase binding sites on μμ-4 DNA for both enzymes as determined bv the heparin technique. φμ-4 DNA is transcribed completely in vitro . In vitro synthesized RNA is, however, not as well translated as other templates in a cell-free E. coli system.

Enzyme synthesisin vitro directed by T7 heteroduplex DNA is observed when the H strand [the strand assuming the higher density in combination with poly(G,U)], carries the wild allele. This is true for enzyme synthesis mediated byE. coli RNA polymerase or by T7 phage RNA polymerase.

The extracellular matrix component hyaluronan is believed to play important roles in various processes of organogenesis, cell migration and cancer. Recognition of and binding to hyaluronan is mediated by cell surface receptors. Three of them, CD44, ICAM-1 and RHAMM (receptor for hyaluronic acid mediated motility), have been identified. A cDNA clone designated RHAMM turned out to possess transforming capacity. Based on this published sequence, we isolated the complete cDNA of the murine gene. The cDNA comprises an open reading frame of 2.3 kb and encodes a 95 kDa protein. The protein carries a hyaluronan binding motif which binds to hyaluronan in vitro but not to heparin or chondroitin sulphate. It is ubiquitously expressed in normal cells and in all tumour cell lines irrespective of their metastatic properties. One tumour cell line, the metastatic Lewis lung carcinoma, expresses a larger 105 kDa variant form of the protein due to a genomic rearrangement. Antibodies raised against the 95 kDa protein were used for subcellular localization studies. The hyaluronan binding protein is not detectable at the cell surface but is rather localized exclusively intracellularly. Clearly, the sequence we have identified encodes a protein with properties substantially different to the RHAMM protein. We tentatively name the protein intracellular hyaluronic acid binding protein, IHABP.

The adenovirus E1A proteins differentially regulate AP-1-responsive genes. Collagenase and stromelysin are repressed by E1A, whereas the expression of c-jun is elevated. Inhibition of collagenase has been found to be exerted through the consensus AP-1 binding site TGAGTCA. Here we show that the distal AP-1 binding site in the c-jun promoter, the jun2TRE (TTACCTCA), is the decisive element of this promoter in mediating the positive response to the 243 amino acid E1A product. In vitro binding studies revealed that, in contrast to the consensus AP-1 site which is preferentially targeted by dimers composed of the Jun and Fos families, the jun2TRE binds heterodimers composed of cJun and ATF-2(-like) proteins. Since stimulation of c-jun transcription is a function of the transforming domain of E1A encoded by conserved region 1, cJun--ATF-2 may be one of the effector factors involved in transformation. The data further suggest that E1A can distinguish between cJun--cJun and cJun--ATF-2 in imposing opposite states of activity.

Protein synthesizing particles from rat liver sedimented more slowly after incubation under protein synthesis conditions. This change paralleled the degree of leucine incorporation which is stimulated by added RNA. Both effects seemed to be specific.

Simian Virus 40 wild type (SV40)-transformed Chinese hamster embryo cells (Co63 1) contain about five viral copies integrated per cell genome. These SV40 sequences were used as endogenous indicator genes to study the response of mammalian cells to radiation at the gene level. An increase in copy number was detected by dispersed cell blotting and Southern analysis in combination with specific DNA hybridization. All types of radiation tested induce a 15- to 25-fold amplification of SV40 sequences without producing intact virus. The amplification is dose dependent and increases with time after irradiation: a maximum effect is observed at Day 3 after a particle or uv exposure and at Day 6 after y-ray exposure. A RBE of 6 can be calculated for a particles if amplification rates at Day 3 are compared. However, when the maximum effect is considered independent of time, no difference between different types of radiation is observed. Southern blots of genomic DNA show that not all integrated SV40 sequences are amplified upon radiation. Amplified sequences are found either in restriction fragments of relatively high molecular weight or in unit size fragments. SV40 amplification is selective in that the amplification of other genes, e.g., of a-actin, dhfr (dihydrofolate reductase), and of two oncogenes of the ras family