Contribution to the study of cytokinin production by phytopathogenic fungi

The excretion of cytokinins into the cultivating medium, which are produced by the phytopathogenic fungiMonilia sp. andCytospora sp. has been investigated. All the isolates of the fungi used in the experiments (Monilia fructicola, Monilia fructigena, isolate 2 and 4,Monilia laxa isolate 3 andCytospora sp. isolate CPL and C₁) have been found to produce cytokinins. The production is increased during the formation of the fructification organs. Among the isolates investigatedMonilia fructigena isolate 4 andCytospora sp. isolate CPL showed the highest production of cytokinins. After chromatographic separation, cytokinin activity was found at R_F 0.7–0.8 values by the biological test as well as by identification according to UV spectra. Application of purified cytokinins produced by the fungi evoked the formation of “green islands” on isolated barley leaves underin vitro conditions.     

Structural studies of the capsular polysaccharide of Klebsiella type 59.

The structure of the capsular polysaccharide from Klebsiella type 59 has been investigated by methylation analysis, a modified Smith-degradation procedure, and uronic acid degradation followed by oxidation and elimination of the substituents of the oxidized residue. The oligomeric fragments produced by these degradative procedures were isolated and characterized. O-Acetyl groups were identified and their position determined. The polysaccharide consists of the following pentasaccharide repeating-unit (dotted lines indicate that only some of the residues carry the O-acetyl substituent). See article.     

Determination in vitro of the rate of protein synthesis and degradation in human-skeletal-muscle tissue.

The present studies were aimed to evaluate the possibility to use a system for estimation in vitro of the biosynthesis and degradation rates of human skeletal muscle protein. A previously characterized human skeletal muscle preparation was used. Amino acids and insulin stimulated significantly the incorporation rate of leucine into proteins. The effect of amino acids was more pronounced than that of insulin. The stimulatory effect of insulin could be decreased by amino acids. Insulin did not influence the tissue uptake or the oxidation rate of leucine. The release of [14C]leucine deriving from degradation of prelabelled skeletal muscle fibre proteins was linear for at least 2.5 h of incubation and optimal with leucine at concentrations beyond 12.5 mmol/1 or in the presence of puromycin in the incubation medium. The rate of the release of radioactivity was significantly inhibited by amino acids and at borderline significance by insulin but not by puromycin. The specific radioactivity in prelabelled proteins decreased significantly in the presence of puromycin suggesting that leucine derived from protein degradation was reutilized in vitro. This reutilization was found to be 9 +/- 1% of leucine released from degradation of proteins in 30 subjects. A statistically significant positive correlation between the cathepsin D activity in human skeletal muscle tissue and the degradative rate of prelabelled muscle proteins in vitro was observed. The results indicate that biosynthesis and degradation of skeletal muscle proteins in this system in vitro were subjected to control mechanisms. It is suggested that the release of radioactivity from prelabelled muscle fibre proteins during incubation probably only reflects the degradation of some rapidly-turning-over proteins.     

Effect of palladium and platinum salts on bacteriophage T4 and its isolated DNA]

The effect of palladium and platinum salts (K2PdCl4, K2PtCl4) on bacteriophage F4 and its isolated DNA in genetic transformation is investigated. Both compounds efficiently inactivated the phage and decreased the transforming activity of donor DNA. The palladium salt exhibited the higher activity. The palladium compounds inhibited the transforming activity of native donor DNA to a greater degree. No difference was observed in the degree of inactivation of the transforming activity of native and denatured DNA under the effect of platinum salt. It is suggested that the difference in the transforming activity of donor DNA treated with the tested compounds reflect the pattern of their interactions with nucleic acids.     

Diadenosine triphosphate splitting by rat liver extracts.

A splitting activity on diadenosine triphosphate has been found in rat liver. One of the products of the cleavage is ADP. A Km of 10 μM has been found. This activity on diadenosine triphosphate seems to be specific as diadenosine tetraphosphate, a nucleotide previously described by others to occur in rat liver at very low concentration, is not a substrate of the reaction. The occurrence of diadenosine triphosphate in rat liver has not been so far reported, but a dinucleoside triphosphate structure has been described at the 5′ end of certain mRNAs. The possibility that this enzymatic activity may be involved in the hydrolysis of diadenosine triphosphate or in the processing of mRNAs is suggested.     

Production in an Actinomyces levoris culture under the action of an actinophage specific to it of variants with stable preservation of its resistance to the phage]

Act. levoris 28, an organism producing levorin was treated with an actinophage virulent to it. Variants of the organism were isolated from the secondary growth of the culture. As a result of lysogenization with the above phage the variants acquired stability to it which was preserved during the further generations. In the previous experiments carried out by the authors the variants isolated from the secondary growth of the culture after its exposure to the same phage lost their stability to the phage as a result of loosing the prophage by it during the subsequent passages. The phage stable variants did not differ from the initial culture either in the activity of levorin or the levorin composition. The phages found in the initial culture 28, and the virulent mutant were identical with respect to the particles morphology and antigenic properties which confirmed their relation.     

Quantitative aspects of hormone-receptor interactions of high affinity: Effect of receptor concentration and measurement of dissociation constants of labeled and unlabeled hormones

It is demonstrated that because of limitations in the magnitude of the specific activity of radiolabeled hormone derivatives, direct binding studies of hormone-receptor interactions of high affinity (10^?9–10^?11 M, depending on whether ³H- or ¹²³I-labeled hormones are used) will be subject to artifactual distortions due to the need to utilize high concentrations of the receptor. If the concentration of the receptor is not ten times lower than the true affinity constant, the apparent dissociation constant obtained from direct concentration binding curves will vary as a linear function of the receptor concentration. In addition, at high receptor concentrations saturability becomes difficult to demonstrate experimentally and the binding data yield apparently non-hyperbolic, sigmoidal curves which can be mistakenly interpreted to depict cooperative interactions. Similar artifacts related to receptor concentration are predicted for measurements of the hormone concentration dependence of biological processes (e.g. activation of adenylate cyclase, transport processes, etc.). Methods for detecting these effects, and correctly measuring affinities for labeled and unlabeled hormones under these conditions, are described. The implications for measuring the binding properties of hormone-receptor interactions are discussed, especially in reference to studies of the comparative analysis of receptor function in altered metabolic states and to studies relating the biological and binding properties of hormones.     

Burkitt's lymphoma - a human tumor model system for immunological studies.

Burkitt's lymphoma occurs mainly in parts of tropical Africa and has attracted the attention of experimental workers due to its epidemiological and clinical features, which indicate a viral etiology and a host immune response to the tumor. As a result of virological studies, Epstein-Barr virus (EBV) DNA has been demonstrated in almost all tested biopsies of African BL. This contrasts to the absence of EBV in all, or almost all, of the non-African Burkitt's lymphoma-like tumors, even though the number of tested tumors in this group is small, and to the lack of EBV in all other types of lymphoma or leukemia. Immunological studies have revealed the presence of antibodies to different EBV-associated antigens in all African patients with Burkitt's lymphoma. However the antibodies are not specific for Burkitt's lymphoma but are found in most adults all over the world, although at lower levels. They cannot therefore serve diagnostic purposes, but they can give prognostic information and occasionally give clues to the mechanisms behind late tumor recurrences, and possibly guide so-called immunotherapy. Burkitt's lymphoma patients contrast to appropriate control groups where some of the persons are anti-EBV seronegative, and this, together with the presence of EBV in Burkitt's lymphoma biopsies and the absence of EBV in other lymphomas, even though the cell type involved may be infectable by EBV in vitro and the tumor may arise in an EBV-carrying person, favors an etiological role in EBV in Burkitt's lymphoma and speaks against the "passenger" hypothesis, according to which EBV is picked up by the Burkitt's lymphoma cell which happens to be particularly suitable for EBV persistence. To explain the geographical distribution, a cofactor, such as certain forms of malaria, has been implied.     

Purification of membrane proteins from Acholeplasma laidlawii by agarose suspension electrophoresis in Tween 20 and polyacrylamide and dextran gel electrophoresis in detergent-free media.

Four of the membrane proteins from Acholeplasma laidlawii that are soluble in the nonionic detergent Tween 20 have been purified by preparative electrophoretic techniques utilizing different supporting media. The last purification step for two of the major proteins was a preparative polyacrylamide gel electrophoresis performed in the absence of any detergent. The proteins were recovered by continuous elution. The purity of the fractions was examined by analytical polyacrylamide gel electrophoresis and crossed immunoelectrophoresis. Two of the minor proteins were purified by dextran gel electrophoresis as the final step, which was also performed in a detergent-free buffer. The separation was followed by scanning the dextran gel in ultraviolet light. The proteins were recovered by slicing the gel and degrading the gel slices with dextranase. The homogeneity of the fractions was checked by electroimmunoassay.     

Derepression and repression of the histidine operon: role of the feedback site of the first enzyme.

Thiazolealanine, a false feedback inhibitor, causes transient repression of the his operon previously derepressed by a severe histidine limitation in strains with a wild-type or feedback-hypersensitive first enzyme but not in feedback-resistant mutants. Since experiments reported here clearly demonstrate that thiazolealanine is not transferred to tRNAHis, it is proposed that this "transient repression" is effected through the interaction of thiazolealanine with the feedback site of the enzyme. Experiments in the presence of rifampin indicate that this thiazolealanine-mediated effect is exerted at the level of translation. We conclude that histidine (free), in addition to forming co-repressor, also represses the operon at the level of translation through feedback interaction with the first enzyme of the pathway (adenosine 5'-triphosphate phosphoribosyltransferase). Rates of derepression in feedback-resistant strains are roughly half of those observed in controls, suggesting a positive role played by a first enzyme with a normal but unoccupied feedback site. Some feedback-resistant mutants, in contrast to the wild type, were unable to exhibit derepression under histidine limitation caused by aminotriazole.