Genetic transformation of somatic cells. VI. Transfer of the trait of the rate of loss of transformant phenotype by means of DNA from cells containing the thymidine kinase gene of the herpes simplex virus

Using dot-hybridization with thymidine kinase gene (tk gene) of Herpes simplex virus type 1 (HSV 1) of DNA preparations obtained from isolated metaphase chromosomes and lysate fractions of metaphase cells, which presumably contain smaller particles compared to metaphase chromosomes, it has been shown that the tk gene of HSV 1 is localized in chromosomes of cells of transformant clones unstable in TK+-phenotype. The DNA isolated from the metaphase chromosomes from cells of transformant clones is 1.5- or 2-fold more efficient in transforming TK-Chinese hamster cells than is the total high molecular weight DNA from the same cells. Upon transformation of TK- cells by the high molecular weight DNA from the tk gene of HSV 1-containing clones, varying in the rate of the loss of TK+-phenotypes, the character "rate of the loss of transformant phenotype" is transferred together with the tk gene of HSV 1 in 22% of cases. Cells of rerevertant clones, produced from TK- subclones of transformant clones, display the rate of the loss of transformant phenotype characteristic of cells of parental TK+-clones. A comparison of the results allows a conclusion that DNA sequences, determining the character "rate of the loss of transformant phenotype", are linked tightly with the transforming DNA proper containing the tk gene of HSV 1, but are not localized inside such a DNA.     

A high affinity topoisomerase I binding sequence is clustered at DNAase I hypersensitive sites in Tetrahymena R-chromatin

Topoisomerase I is associated with DNAase I hypersensitive sites in the nontranscribed spacers flanking the rRNA genes in Tetrahymena thermophila. The endogenous topoisomerase I introduces site and strand specific single-strand cleavages in the rDNA spacers in situ. The cleavages occur base specifically within a hexadecameric sequence element present in two or three direct repeats at the hypersensitive sites. The sequence specificity and polarity of the cleavage reaction are identical when the enzyme is reacted with naked rDNA, indicating that the repetitive element functions as a high-affinity topoisomerase I attraction site in the r-chromatin. The biological mechanism associated with this phenomenon appears to be widespread among eukaryotes, since the topoisomerase I recognition sequence is conserved in the rDNA spacers of phylogenetically remote organisms, such as fungi, slime molds, ciliates, and insects.     

Successful transfer of embryos recovered from a cow with chronic cervicitis of mixed aerobic and anaerobic bacterial aetiology a case report

An imported 13-year-old Simmentaler cow was presented with severe purulent cervicitis and endometritis from which Corynebacterium pyogenes , Fusobacterium necrophorum , Bacteroides melaninogenicus and an untyped Clostridium perfringens were isolated. The endometritis responded to treatment but the cervix did not and remained fibrous, continuously patent and purulent. Natural pregnancy was considered unlikely and the cow was prepared as an embryo transfer donor. Eight embryos were recovered and six transferred, resulting in five confirmed pregnancies and four live births.     

Possible role of hypoxia and lipid peroxidation in the development of a neurosis-like condition in the rat

The chronic emotional pain stress resulting in a development of neurosis-like state in rats induced an increase of arterial pressure and change of the cardiac rate dynamics under the conditions of functional load. An increase of cardiac mass was also seen without change of masses of the thymus, adrenal glands and the spleen. The rise of activity of cytochromeoxidase and activation of peroxide lipide oxidation (by malonate dialdehyde level) were observed in the cerebral cortex and the hippocampus of neurotized rats. Injection of antioxidant F-801 before each emotional pain stress trial prevented vegetative disturbances, cardiac hypertrophy, and increase of oxidative activity in the brain. The role of peroxide lipide oxidation and that of the factor of hypoxia in development of disturbances caused by neurotization were discussed.     

Study of the effect of β-1,3-glucanase fromBasidiomycete QM 806 on yeast extract production

Summary Cell free supernatants, containing -1,3-glucanase fromBasidiomycete QM 806, dramatically augmented the effect of papain on yeast autolysis. This enables the process time to be significantly reduced and/or the yield of extract to be substantially increased.     

Vitamin A effects on UMR 106 osteosarcoma cells are not mediated by specific cytosolic receptors.

Retinol and retinoic acid at 20 microM altered cell morphology and inhibited cell proliferation of UMR 106 osteosarcoma cells in culture. No specific cytosolic binding proteins for retinol could be detected.     

Effects of combined administration ofl-tryptophan and tricyclic antidepressants on α2- and β-adrenoceptors and monoamine levels in rat brain

In order to test whether co-administration of a serotonin precursor with antidepressant drugs could potentiate the effects of the antidepressants on monoamines or adrenoceptors in rat brain,l-tryptophan (20 mg/kg) was administered to rats daily for 7 or 15 days, either alone or in combination with desipramine (10 mg/ kg) or amitriptyline (10 mg/kg). After treatment withl-tryptophan for 7 days, increases were observed in rat hypothalamic and frontal cortex 5-hydroxy-3-indoleacetic acid levels as well as in hypothalamic dopamine and nucleus accumbens 3,4-dihydroxyphenylacetic acid levels. After 15 days, hippocampal -adrenoceptor density was found to be decreased. There was no evidence of potentiation of desipramine or amitriptyline action whenl-tryptophan was administered in combination with the antidepressants. On the contrary, the antidepressants appeared to interact withl-tryptophan to reduce its effects.     

Deposition of type X collagen in the cartilage extracellular matrix

In cultured chick embryo chondrocytes, type X collagen is preferentially deposited in the extracellular matrix, the ratio between type II and type X collagen being about 5 times higher in the culture medium than in the cell layer. When the newly synthesized collagens deposited in slices from the epiphyseal cartilage of 17-day-old embryo tibiae were isolated, type X collagen was always the major species. In agreement with this result the mRNA for type X collagen was the predominant mRNA species purified from the same tissue. When the total collagen (unlabeled) deposited in the epiphyseal cartilage was analyzed, it was observed that type X collagen represented only 1/15 of the type II collagen recovered in the same preparation. The possible explanations for these differences are discussed.     

DNA single-strand breaks,double-strand breaks,and crosslinks in rat testicular germ cells: Measurements of their formation and repair by alkaline and neutral filter elution

This work describes a neutral and alkaline elution method for measuring DNA single-strand breaks (SSBs), DNA double-strand breaks (DSBs), and DNA-DNA crosslinks in rat testicular germ cells after treatments in vivo or in vitro with both chemical mutagens and gamma-irradiation. The methods depend upon the isolation of testicular germ cells by collagenase and trypsin digestion, followed by filtration and centrifugation. ¹³⁷Cs irradiation induced both DNA SSBs and DSBs in germ cells held on ice in vitro. Irradiation of the whole animal indicated that both types of DNA breaks are induced in vivo and can be repaired. A number of germ cell mutagens induced either DNA SSBs, DSBs, or cross-links after in vivo and in vitro dosing. These chemicals included methyl methane sulfonate, ethyl methane sulfonate, ethyl nitrosurea, dibromochlorpropane, ethylene dibromide, triethylene melamine, and mitomycin C. These results suggest that the blood-testes barrier is relatively ineffective for these mutagens, which may explain in part their in vivo mutagenic potency.This assay should be a useful screen for detecting chemical attack upon male germ-cell DNA and thus, it should help in the assessment of the mutagenic risk of chemicals. In addition, this approach can be used to study the processes of SSB, DSB, and crosslink repair in DNA of male germ cells, either from all stages or specific stages of development.Abbreviations DBCP dibromochlorpropane - DSB(s) DNA double-strand break(s) - EDB ethylene dibromide - EMS ethyl methane sulfonate - ENU ethyl nitrosurea - MC mitomycin C - MMS methyl methane sulfonate - SDS sodium dodecyl sulfate - SSB (s) DNA single-strand break(s) - TEM triethylene melamine - UDS unscheduled DNA synthesis