Promoters and processing sites within the transforming region of bovine papillomavirus type 1.

The mRNAs present in bovine papillomavirus type 1 (BPV-1)-transformed C127 cells were studied by primer extension. The results show that two internal promoters are present in the E region of BPV-1 in addition to the previously identified promoter at coordinate 1 (H. Ahola, A. Stenlund, J. Moreno-López, and U. Pettersson, Nucleic Acids Res. 11:2639-2650, 1983). One, located at coordinate 31, generated a set of mRNAs with heterogeneous 5' ends, which may encode the major transforming protein of BPV-1, the E5 protein. The second promoter, which is located at coordinate 39, generates colinear mRNAs which encode either the E4 protein or a truncated form of the E2 protein. Unlike the cottontail rabbit papillomavirus (O. Danos, E. Georges, G. Orth, and M. Yaniv, J. Virol. 53:735-741, 1985), BPV-1 appears to lack a separate promoter for expression of the E7 protein. The major splice sites in the transforming region (E region) of the BPV-1 genome were also identified by nucleotide sequence analysis.     

European elk papillomavirus: characterization of the genome, induction of tumors in animals, and transformation in vitro.

The European elk papillomavirus (EEPV) genome was cloned in the BamHI cleavage site of the pBR322 vector. The cloned genome was used for construction of a physical map, employing restriction endonucleases BamHI, BglII, HindIII, PvuII, SacI, and XhoI. The sequence homology between the EEPV and bovine papillomavirus type 1 genomes was elucidated by performing hybridizations in different concentrations of formamide. Sequence homology could only be revealed under less stringent conditions, i.e., Tm - 43 degrees C. Nucleotide sequence information was also collected from the regions which lie adjacent to the three HindIII sites that are present in the EEPV genome. The results made it possible to align the EEPV and bovine papillomavirus type 1 genomes. Transformation by EEPV was demonstrated with the C127 mouse cell line, and fibrosarcomas were induced in young hamsters after subcutaneous injection. The transformed cells and the tumors contain multiple, nonintegrated copies of the EEPV genome. Virus particles could not be detected either in tumors or in transformed cells.     

Environmental correlates of annual survival differ between two ecologically similar and congeneric owls

Understanding how survival is affected by the environment is essential to gain insight into population dynamics and the evolution of life‐history traits as well as to identify environmental selection pressures. However, we still have little understanding of the relative effect of different environmental factors and their interactions on demographic traits and population dynamics. Here we used two long‐term, individual‐based datasets on Tawny Owl Strix aluco (1981–2010) and Ural Owl S. uralensis (1986–2010) to undertake capture‐mark‐recapture analysis of annual survival of adult females in response to three biologically meaningful environmental variables and their two‐way interactions. Despite the similar ecology of these two species, their survival was associated with different and uncorrelated environmental drivers. The main correlate of Tawny Owl survival was an inverse association with snow depth (winter severity). For Ural Owl, high food (vole) abundance improved survival during years with deep snow, but was less important during years with little snow. In addition, Ural Owl survival was strongly density‐dependent, whereas Tawny Owl survival was not. Our findings advise caution in extrapolating demographic inferences from one species to another, even when they are very closely related and ecologically similar. Analyses including only one or few potential environmental drivers of a species' survival may lead to incomplete conclusions because survival may be affected by several factors and their interactions.     

Brown tawny owls moult more flight feathers than grey ones

The mechanisms by which melanin‐based colour polymorphism can evolve and be maintained in wild populations are poorly known. Theory predicts that colour morphs have differential sensitivity to environmental conditions. Recently it has been proposed that colour polymorphism covaries genetically with intrinsic and behavioural properties. Plumage moult is a costly and crucial somatic maintenance function in birds. We used a long‐term data set consisting of 761 observations on 307 individuals captured between 1985 and 2010 to examine differences in partial flight feather moult between grey (pale) and brown (pheomelanic dark) colour morphs of the tawny owl. We find that the brown morph consistently moult more primary flight feathers than the grey morph whereas there is no clear difference between colour morphs in the moulting of secondary feathers. Contrary to expectations, the difference in the number of moulted flight feathers between the morphs was independent of environmental conditions, as quantified by the abundance of prey. We discuss the potential physiological and behavioural causes for and costs of the observed difference in maintenance functions between colour morphs.     

Versatile Trans-Replication Systems for Chikungunya Virus Allow Functional Analysis and Tagging of Every Replicase Protein

Chikungunya virus (CHIKV; genus Alphavirus, family Togaviridae) has recently caused several major outbreaks affecting millions of people. There are no licensed vaccines or antivirals, and the knowledge of the molecular biology of CHIKV, crucial for development of efficient antiviral strategies, remains fragmentary. CHIKV has a 12 kb positive-strand RNA genome, which is translated to yield a nonstructural (ns) or replicase polyprotein. CHIKV structural proteins are expressed from a subgenomic RNA synthesized in infected cells. Here we have developed CHIKV trans-replication systems, where replicase expression and RNA replication are uncoupled. Bacteriophage T7 RNA polymerase or cellular RNA polymerase II were used for production of mRNAs for CHIKV ns polyprotein and template RNAs, which are recognized by CHIKV replicase and encode for reporter proteins. CHIKV replicase efficiently amplified such RNA templates and synthesized large amounts of subgenomic RNA in several cell lines. This system was used to create tagged versions of ns proteins including nsP1 fused with enhanced green fluorescent protein and nsP4 with an immunological tag. Analysis of these constructs and a matching set of replicon vectors revealed that the replicases containing tagged ns proteins were functional and maintained their subcellular localizations. When cells were co-transfected with constructs expressing template RNA and wild type or tagged versions of CHIKV replicases, formation of characteristic replicase complexes (spherules) was observed. Analysis of mutations associated with noncytotoxic phenotype in CHIKV replicons showed that a low level of RNA replication is not a pre-requisite for reduced cytotoxicity. The CHIKV trans-replicase does not suffer from genetic instability and represents an efficient, sensitive and reliable tool for studies of different aspects of CHIKV RNA replication process.     

Coenzyme Q10 supplementation and recovery from ischemia in senescent rat myocardium

Many studies have suggested that parenteral administration of coenzyme Q10 (Q10) protects the myocardium of young experimental animals from post-ischemic reperfusion injury. Although parenteral administration, in contrast to per os supplementation, seems to elevate coenzyme Q concentrations in heart tissue, it is not suitable for prophylactic use. In addition, the incidence of ischemic events is greatest in older age. We studied the effect of Q10 supplementation on myocardial postischemic recovery in 18-month-old Wistar rats. The treated group (n=9) received 10 mg/kg/day of Q10 for 8 weeks in their chow while the normal chow of the control group (n=9) contained less than 0.5 mg/kg/day of Q10. The treatment clearly elevated plasma Q10 concentration (286 +/- 25 micromol/l and 48 +/- 30 micromol/l, treated and controls, respectively, p<0.0001) but neither Q9 nor Q10 concentrations in heart tissue were affected by the supplementation. The isolated perfused hearts were subjected to 20 minutes of ischemia and 30 minutes of reperfusion. The preischemic values of developed pressure (DP) but not contractility (+DP/delta t) and relaxation (-DP/delta t) were improved by Q10 supplementation (p=0.034, p=0.057 and p=0.13, respectively) while in postischemic recovery no differences were observed between the groups (p>0.05 at all time points). Also, in myocardial flow, myocardial oxygen consumption (MVO2) and myocardial aerobic efficiency (DP/MVO2) the groups did not differ at any time points. Although dietary Q10 supplementation clearly elevated plasma Q10 concentrations in senescent rats, the coenzyme Q contents in heart tissue and myocardial recovery from ischemia were not affected. However, it is possible that the site of action for the reported beneficial effects of Q10 is in the coronary endothelium rather than myocardium itself.     

Involvement of tyrosine kinases on cyclooxygenase expression and prostaglandin E2 production in human gingival fibroblasts stimulated with interleukin-1beta and epidermal growth factor

The purpose of the present study was to investigate the involvement of cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), and tyrosine kinase on prostaglandin E2 (PGE2) production in human gingival fibroblasts stimulated by interleukin-1beta (IL-1beta) and/or epidermal growth factor (EGF). The cytokine IL-1beta and to a lesser extent EGF, enhanced COX-2 mRNA levels in gingival fibroblasts. Simultaneous treatment with EGF and IL-1beta resulted in enhanced COX-2 mRNA levels accompanied by a synergistic stimulation of PGE2 biosynthesis compared to the cells treated with IL-1beta or EGF alone. Neither IL-1beta EGF nor the combination of IL-1beta and EGF enhanced COX-1 mRNA levels in gingival fibroblasts. The tyrosine kinase inhibitors, Herbimycin A and PD 153035 hydrochloride, reduced COX-2 mRNA levels as well as PGE2 production induced by IL-1beta or the combination of IL-1beta and EGF whereas COX-1 mRNA levels were not affected. Furthermore, the COX-2 specific inhibitor, NS-398, abolished the PGE2 production induced by IL-1beta, EGF, or the combination. These results indicate that the synergy between IL-1beta and EGF on PGE2 production is due to an enhanced gene expression of COX-2 and that tyrosine kinase(s) are involved in the signal transduction of COX-2 in gingival fibroblasts.     

A prevalent POLG CAG microsatellite length allele in humans and African great apes

The human nuclear gene for the catalytic subunit of mitochondrial DNA polymerase (POLG) contains within its coding region a CAG microsatellite encoding a polyglutamine repeat. Previous studies demonstrated an association between length variation at this repeat and male infertility, suggesting a mechanism whereby the prevalent (CAG)₁₀ allele, which occurs at a frequency of >80% in different populations, could be maintained by selection. Sequence analysis of the POLG CAG microsatellite region of more than 1000 human chromosomes reveals that virtually all allelic variation at the locus is accounted for by length variation of the CAG repeat. Analysis of POLG from African great apes shows that a prevalent length allele is present in each species, although its exact length is species-specific. In common chimpanzee (Pan troglodytes) a number of different sequence variants contribute to the prevalent length allele, strongly supporting the idea that the length of the POLG microsatellite region, rather than its exact nucleotide or amino acid sequence, is what is maintained. Analysis of POLG in other primates indicates that the repeat has expanded from a shorter, glutamine-rich sequence, present in the common ancestor of Old and New World monkeys.