Interaction of poly(ADP-ribose)polymerase with DNA polymerase α

Homogeneously purified poly(ADP-ribose) polymerase (PARP) specifically stimulated the activity of immunoaffinity-purified calf or human DNA polymerase by about 6 to 60-fold. Apparently, poly(ADP-ribosyl)ation of DNA polymerase was not necessary for the stimulation. The effects of PARP on DNA polymerase were biphasic: at very low concentrations of DNA, it rather inhibited its activity, whereas, at higher DNA concentrations, PARP greatly stimulated it. The autopoly(ADP-ribosyl)ation of PARP suppressed both its stimulatory and inhibitory effects. By immunoprecipitation with an anti-DNA polymerase antibody, it was clearly shown that PARP may be physically associated with DNA polymerase . Stimulation of DNA polymerase may be attributed to the physical association between the two, rather than to the DNA-binding capacity of PARP, since the PARP fragment containing only the DNA binding domain showed little stimulatory activity. The existence of PARP-DNA polymerase complexes were also detected in crude extracts of calf thymus.     

Molecular biology and systematics of an extinct Lemur:Pachylemur insignis

The systematic position of the extinctPachylemur insignis has been controversial: some authors consideredPachylemur as a lemur, whereas others viewed it closer toVarecia. Its classification in the genusLemur orVarecia thus remained an open question. DNA extraction from subfossil bones, using a non-destructive method, allowed us to obtain enough material to make a Southern blot. The hybridization ofPachylemur withEulemur fulvus, Lemur catta, andVarecia variegata highly repeated DNA probes showed that only theVarecia probe gave a positive signal on hybridization on thePachylemur blot. These results indicate thatPachylemur must be considered closer to the genusVarecia than toEulemur andLemur.     

The methylation status of DNA derived from potato plants recovered from slow growth

The in vitro conservation of potato using tissue culture medium supplemented with the growth retardant mannitol causes morphological changes in the propagated material. These culture conditions seem to have an affect on the DNA extracted from the regenerated plants, when it is digested by the methylation sensitive restriction enzymes Hpa II/Msp I and Eco RII/Bst NI, compared to the control material. In most of these plants, there appears to be preferential methylation of nuclear domains that contain Eco RII/Bst NI recognition sites in contrast to those that contain Hpa II/Msp I sites. The refractory nature of the isolated DNA to these restriction enzymes was attributed to hypermethylation of genomic DNA and the ribosomal RNA genes. These findings indicate that methylation of DNA sequences may be an adaptive response to conditions of high osmotic stress. The importance of these results for the conservation of potato germplasm and international exchange is discussed.     

Molecular phylogeny of the Praomys complex (Rodentia: Murinae): a study based on DNA/DNA hybridization experiments.

Within the Murinae (Muridae: Rodentia), the African rats of the Praomys group, whose systematics has been studied through different approaches, have raised numerous taxonomic problems. Different taxa related to Praomys have successively been described, among which Mastomys, Myomys and Hylomyscus were considered either as separate genera or subgenera of Praomys. In order to clarify the relationships within the Praomys group, we conducted a series of DNA/DNA hybridization experiments involving different species of Praomys, Mastomys, Myomys and Hylomyscus plus other Murinae and a Cricetomyinae. This study indicates that the Praomys complex is a monophyletic entity clearly separated from the other African and Asian Murinae. If Mastomys and Hylomyscus appeared to be independent genera, the taxonomic situation of Praomys and Myomys is more difficult to ascertain. Indeed, Praomys tullbergi appears more closely related to Myomys daltoni than to another species of Praomys , namely P. jacksoni , suggesting paraphyly for Praomys. Furthermore, P. jacksoni is as distant from P. tullbergi as from any species of Mastomys. Additional species of Praomys and, especially, of Myomys , are needed for reaching a definitive conclusion on these latter taxa. The Praomys group is more related to Mus than to Rattus. To calibrate our molecular distances with geological time, we used a dating of 10 Myr for the Musi Rattus dichotomy. The inferred rate of molecular evolution suggests a dating of c. 8 Myr for the separation of the Praomys group from the Mus lineage.     

Analysis of DNA polymerase activity in Petunia protoplasts treated with clastogenic agents

Clastogenic agents, i.e. agents that can induce chromosome or DNA breakage, have been shown to enhance the rale of direct gene transfer to protoplasts. The effect was analysed at the enzymatic level using protoplast homogenates as well as intact protoplasts. For that purpose existing procedures were modified to enable measurement of DNA polymerase in vivo. In the system used, external DNA was able to enter the cells without the addition of membrane-permeabilizing compounds. When comparing total DNA polymerase activity of protoplasts irradiated with X-rays or UV-light with that of untreated cells we did not observe significant differences. Incubation of protoplasts with high doses of bleomycin affected total DNA polymerase activity negatively. but dideoxythymidine triphosphate-sensitive activity was not influenced. We conclude that the DNA strand-breaks induced by low doses of X-rays. UV-light or bleomycin do not increase the total or the repair-DNA polymerase activity and. therefore. that the increase in the transformation rates after DNA strand-breaking is not preceded by enhanced DNA polymerase activity.     

The evolutionary origin of the 35 kb circular DNA of Plasmodium falciparum: new evidence supports a possible rhodophyte ancestry

In common with other Apicomplexan parasites, Plasmodium falciparum carries two extrachromosomal DNAs, one of which, the 6 kb element, is undoubtedly mitochondrial. The second, generally referred to as the 35 kb circle, is of unknown provenance, but the nature and organization of its genetic content makes a mitochondrial association unlikely and the molecule has features reminiscent of plastid genomes. We now report the occurrence on the circle of an open reading frame specifying a predicted 470 amino acid protein that shares more than 50% identity with a gene currently known only on the plastome of red algae. This high degree of conservation confirms the 35 kb circle's plastid ancestry, and we speculate that it may have originated from the rhodoplast of an ancient red algal endosymbiont in the progenitor of the Apicomplexa.     

How does the G protein,Gi2, transduce mitogenic signals?

Serpentine receptors coupled to the heterotrimeric G protein, G_i2, are capable of stimulating DNA synthesis in a variety of cell types. A common feature of the G_i2-coupled stimulation of DNA synthesis is the activation of the mitogen-activated protein kinases (MAPKs). The regulation of MAPK activation by the G_i2-coupled thrombin and acetylcholine muscarinic M₂ receptors occurs by a sequential activation of a network of protein kinases. The MAPK kinase (MEK) which phosphorylates and activates MAPK is also activated by phosphorylation. MEK is phosphorylated and activated by either Raf or MEK kinase (MEKK). Thus, Raf and MEKK converge at MEK to regulate MAPK. G_i2-coupled receptors are capable of activating MEK and MAPK by Raf-dependent and Raf-independent mechanisms. Pertussis toxin catalyzed ADP-ribosylation of α_i2 inhibits both the Raf-dependent and-independent pathways activated by G_i2-coupled receptors. The Raf-dependent pathway involves Ras activation, while the Raf-independent activation of MEK and MAPK does not involve Ras. The Raf-independent activation of MEK and MAPK most likely involves the activation of MEKK. The vertebrate MEKK is homologous to the Ste11 and Byr2 protein kinases in the yeast Saccharomyces cerevisiae and Schizosaccharomyces pombe, respectively. The yeast Ste11 and Byr2 protein kinases are involved in signal transduction cascades initiated by pheromone receptors having a 7 membrane spanning serpentine structure coupled to G proteins. MEKK appears to be conserved in the regulation of G protein-coupled signal pathways in yeast and vertebrates. Raf represents a divergence in vertebrates from the yeast pheromone-responsive protein kinase system. Defining MEKK and Raf as a divergence in the MAPK regulatory network provides a mechanism for differential regulation of this system by G_i2-coupled receptors as well as other receptor systems, including the tyrosine kinases.