Simian Virus 40 Transcription in Productively Infected and Transformed Cells

Several independent cell lines transformed by simian virus 40 carry a species of viral RNA of 900,000 to 1,000,000 daltons. A viral RNA species of similar size is found early in the lytic cycle. Late in the viral lytic cycle, two prominent viral RNA species of about 600,000 and 900,000 daltons are seen. The larger late species shares nucleotide sequences with, and is less stable than, the smaller. These RNA species are located in the cytoplasm of the infected cell. The regions of the viral genome coding for these RNA species are mapped by hybridization of lytic RNA species to fragments of the genome produced by cleavage with Haemophilus aegyptius endonuclease.     

Regulation by recombinant interleukin-2 of protective immunity against recurrent herpes simplex virus type 2 genital infection in guinea pigs.

The goal of our study was to determine whether recombinant interleukin-2 (rIL-2) could modify the recurrence pattern of chronic herpes simplex virus type 2 (HSV-2) genital infection in guinea pigs. Animals that developed symptomatic acute HSV-2 infection were distributed at 14 days after viral inoculation into several treatment groups, which were similar with respect to the severity of acute disease. Three rIL-2 dosages administered for 4 weeks in daily subcutaneous injections were tested in this study: 5 X 10(3), 5 X 10(4), and 2.5 X 10(5) U. Daily observations of the animals showed a significant decrease of the incidence of new recurrent lesions with the use of 5 X 10(4) U of rIL-2 (rate of recurrence, 0.08, compared with 0.21 in untreated controls), whereas the other rIL-2 regimens did not affect the overall rate of recurrence. Weekly analysis of recurrences showed that treatment with 5 X 10(4) U of rIL-2 was effective only during the first 3 weeks of use and that 2.5 X 10(5) U of rIL-2 markedly decreased the rate of recurrence in the first week of treatment but not in subsequent weeks. The loss of clinical protection in both groups coincided with the production of neutralizing antibodies to rIL-2. The immune mechanisms possibly involved in the protective effect of rIL-2 in chronic HSV-2 disease were further investigated. Production of gamma interferon correlated well with clinical protection, and circulating levels dropped at the time when neutralizing antibodies to rIL-2 developed. Nonspecific cytotoxicity represented by natural killer cell and lymphokine-activated killer cell activities was also increased in the treated guinea pigs. Antibody titers and lymphocyte proliferation to herpes simplex antigen were similar in rIL-2 and placebo recipients. Finally, we found that the rIL-2-induced immune stimulation was as protective against recurrent HSV-2 disease in guinea pigs as the viral suppression achieved with acyclovir. However, the biological activity of both drugs was not additive when they were coadministered.     

Effect of the bacterial DNA gyrase inhibitors, novobiocin, nalidixic acid, and oxolinic acid, on oxidative phosphorylation

When incubated with isolated intact rat liver mitochondria, novobiocin and nalidixic acid act as uncouplers of oxidative phosphorylation; they stimulate oxygen uptake and inhibit ATP synthesis. Novobiocin is about as powerful an uncoupler as is 2,4-dinitrophenol, nalidixic acid is somewhat less powerful, and oxolinic acid exerts no inhibition whatsoever at the concentrations used. The three inhibitors are without effect on oxidative phosphorylation in Escherichia coli nor does novobiocin affect this process in a novobiocin-permeable mutant of yeast. While it would appear that oxolinic acid may be a relatively specific tool for the manipulation of the superhelicity of DNA in complex systems such as mammalian mitochondria and intact mammalian cells, the specificity of each of these inhibitors may depend upon the particular conditions and species used and such experiments require adequate controls on oxidative phosphorylation.     

p185, a product of the neu proto-oncogene, is a receptorlike protein associated with tyrosine kinase activity.

The neu oncogene was originally identified in cell lines derived from rat neuroectodermal tumors. neu is related to but distinct from the c-erbB gene, which encodes the epidermal growth factor (EGF) receptor. neu encodes a protein, designated p185, that is serologically related to the EGF receptor. Identification of the normal homolog of p185 encoded by the neu proto-oncogene enabled us to compare the product of the neu proto-oncogene with the mutated version specified by the neu oncogene and with the EGF receptor. The normal form of p185 was structurally similar to its transforming counterpart, indicating that activation of the neu oncogene did not cause major structural alterations in the gene product. Both normal and transforming forms of p185 were associated with tyrosine kinase activity, supporting the idea that normal p185 functions as a growth factor receptor. p185 differed both structurally and functionally from the EGF receptor. p185 and the EGF receptor had distinct electrophoretic mobilities when synthesized under normal culture conditions or in the presence of tunicamycin. EGF did not stimulate increased turnover of p185 and did not bind quantitatively to p185. A number of other growth factors failed to stimulate degradation of p185 or tyrosine phosphorylation of p185 and are therefore unlikely to be ligands for p185.     

Motility of isolated mammalian gastric fundus

Gastric fundus strips (GFS) of rats, guinea-pigs, rabbits, cats and humans, but not of dogs, show spontaneous phasic contractions in vitro. Rabbit, cat, dog and human GFS exhibit dose-dependent responses to acetylcholine (ACh), histamine (His), serotonin (5-HT) and KCl. Guinea-pig drug-induced responses are not dose-dependent. Rat GFS does not respond to His. ED50S for KCl are always higher than for other agonists. ED50S for 5-HT are lower than for other drugs in rats and cats, but not different in other species. Maximal GFS contractions induced by ACh are usually higher than those induced by other drugs.     

The HL-60 transforming sequence: a ras oncogene coexisting with altered myc genes in hematopoietic tumors

The oncogene of the HL-60 human promyelocytic leukemia cell line has been passed serially through NIH/3T3 mouse fibroblasts. Oncogene-specific probes prepared from the resulting tertiary transfectants by molecular cloning have been used to show that loss of the transfected oncogene from NIH/3T3 cells correlates with reversion to nontransformed morphology. Analysis of cells transfected by the oncogenes of other tumors and tumor cell lines indicates that the transforming gene of the HL-60 leukemia cell line is closely related to oncogenes of a Burkitt's lymphoma, an acute myelogenous leukemia, an adenocarcinoma of the colon, a neuroblastoma, and two sarcomas. This oncogene is distantly related to the viral oncogenes of Kirsten and Harvey sarcoma viruses. It has been termed N-ras. The active N-ras oncogene coexists with altered versions of the myc oncogene in the HL-60 and AW Ramos human tumors. This suggests a multistep mechanism involving both ras and myc genes in the creation of these tumors.     

Mechanisms of gentamicin-induced dysfunction of renal cortical mitochondria: II. Effects on mitochondrial monovalent cation transport

Mitochondrial swelling techniques were used to evaluate the effects of the aminoglycoside antibiotic gentamicin on renal cortical mitochondrial monovalent cation permeability. Gentamicin behaved like EDTA to enhance energy-dependent Na⁺- and K⁺-acetate uptake with a relatively greater effect on Na⁺-acetate uptake. Mg²⁺ prevented and reversed the effects of both EDTA and gentamicin. Neither agent affected energy-independent uptake of Na⁺ and K⁺-acetate. Gentamicin did not enhance energy-independent uptake of K⁺- and Na⁺-nitrate. Gentamicin enhanced energy-dependent swelling in a chloride- and phosphate-containing medium as a function of the medium Na⁺ and K⁺ concentration. This effect occurred simultaneously with gentamicin-induced stimulation of State 4 respiration and was blocked by Mg²⁺. Gentamicin did not affect phosphate transport. The results are taken to indicate a specific action of gentamicin to enhance mitochondrial monovalent cation permeability at an Mg²⁺-sensitive site and it is proposed that this accounts for the effects of gentamicin on mitochondrial respiration.     

Isolation and Characterization of Simian Virus 40 Ribonucleic Acid

Deoxyribonucleic acid-ribonucleic acid (RNA) hybridization in formamide was used to isolate simian virus 40-specific RNA. Early in the lytic cycle, a 19S viral RNA species was observed. Late in the lytic cycle, 16S and 19S viral species were found. The 16S and 19S species of viral RNA were localized in the cytoplasm. High-molecular-weight heterogeneous RNA, containing viral sequences, was isolated from the nuclear fraction of infected cells late in the lytic cycle. This RNA may contain non-viral sequences linked to viral sequences. The formamide hybridization technique can be used to isolate intact late lytic viral RNA which is at least 99% pure.