ATP(GTP)-dependent conversion of MVM parvovirus single-stranded DNA to its replicative form by a purified 10 S species of mouse DNA polymerase alpha

A species of DNA polymerase alpha that is active in the ATP(GTP)-dependent conversion of MVM parvovirus single-stranded linear DNA to the duplex replicative form has been purified 4300-fold from Ehrlich ascites mouse tumour cells. The single-stranded----replicative form activity is maintained throughout ammonium sulfate precipitation, DEAE-cellulose, phosphocellulose and hydroxyapatite column chromatography and glycerol gradient sedimentation. Polypeptides with Mr = 230 000, 220 000, 183 000, 157 000, 125 000, 70 000, 65 000, 62 000, 57 000, 53 000 and 48 000 copurify with the single-stranded----replicative form activity, which sediments at approx. 10 S. The Mr = 183 000, 157 000 and 125 000 polypeptides exhibit catalytic activity when assayed in situ following SDS-polyacrylamide gel electrophoresis. The 10 S form of DNA polymerase alpha is functionally distinguishable from an 8.4 S form of the enzyme obtained from the same cells on the basis of single-stranded----replicative form activity. The single-stranded----replicative form activity of the 10 S enzyme is stable at 22 degrees C for up to 3 h, but exhibits a half life of only 5 min at 45 degrees C.     

Conditions of formation of the heparin-fibronectin-collagen complex and the effect of plasmin

The formation and composition of the insoluble heparin-fibronectin-collagen complex and its degradation by proteolysis was investigated. At fixed concentrations of the other molecular components of the complex, the maximal rate of complex formation, measured turbidimetrically, was reached at a concentration of 4 microM heparin and 0.9 microM collagen, while the rate of complex formation was linearly related to concentrations of fibronectin as high as 3 microM. Heparin was incorporated into the complex in a saturable manner, and was released in active anticoagulant form by plasmin but not by urokinase. The complex formation was inhibited by 5 mM calcium or 250 mM NaCl as well as by polybrene or spermin. It is suggested that fibronectin binds both heparin and collagen cooperatively to form an insoluble ternary complex of the extracellular matrix.     

Time-dependent subpopulation induction in heterogeneous tumors

Time-dependent induction of clonal heterogeneity in the neoplastic micro-environment is analysed within the context of a competitive ecology. A model that describes a constant source for clonal emergence was analysed by Michelsonet al. (1987) as an extension of a model proposed by Jansson and Revesz (1974). The extended model has been termed the JRE Model. This paper extends these analyses to time-dependent emergence rates which may represent induction in the presence of a cytotoxic agent. If the analysis is constrained to the tumor micro-environment, and if the emergent subpopulation is drug resistant, then the model may describe the induction and emergence of drug resistant subclones in a growing neoplasm. Asymptotic closed form solutions are derived for a class of emergence rate functions which decay asymptotically to a constant mutation rate. This underlying mutation rate may represent spontaneous mutation to the resistant phenotype, and has been analysed stochastically (Coldmanet al., 1985). The asymptotic solutions to the time-dependent model approach the steady state solution for the JRE Model which represents the dynamics observed in the presence of a constant, spontaneous mutation rate. The clinical and biological implications of these results are discussed. Research support provided in part by Hungarian National Foundation for Scientific Research Grant No. 6032/6319 and ACS Grant IN45-Z and ACS PDT 243B.     

Characterization of acute and latent herpes simplex virus infection of dorsal root ganglia in rats.

The characteristics of HSV type-1 infection following subcutaneous inoculation in the dorsum of one hind paw of Sprague-Dawley rats were studied to determine whether infection in rats might more closely parallel the infection in man than is seen in other animals. The serologic and virologic characteristics of acute and latent ganglion infection conformed to those of human infection. Immunohistochemical studies suggested that sensory ganglion infection arose via centripetal axonal migration of virus as is hypothesized in man. In rat, small type B neuronal cell bodies appeared central to the maintenance of latent infection and reactivation observed during cocultivation of lumbar ganglia. Acute and latent lumbar sensory ganglion infection in rats after subcutaneous hind paw injection of HSV-1 appears to be another suitable model of this infection in man.     

Cis-analysis of a seed protein gene promoter: the conservative RY repeat CATGCATG within the legumin box is essential for tissue-specific expression of a legumin gene

A 2.4 kb fragment containing the 5'-flanking region and the 5'-noncoding sequence of the Vicia faba legumin gene LeB4 mediates high level seed-specific expression in transgenic tobacco plants. Deleted derivatives of this legumin upstream sequence were fused to the npt-II reporter gene to determine the tissue-specific activity of the chimeric constructs in stably transformed tobacco plants. The results indicate the presence of positive regulatory, enhancer-like cis elements within 566 bp of the upstream sequence. Most importantly, however, these elements are only fully functional in conjunction with the core motif CATGCATG of the legumin box around position -95, since destruction of the motif by a 6 bp deletion in an otherwise intact 2.4 kb upstream sequence drastically reduces expression in seeds. At the same time, low level expression in leaves is observed. The occurrence of similar CATGCATG consensus cis elements with alternating purine and pyrimidine base pairs in front of several other plant genes suggests a functional role of the motif in a wider range of plant promoters.