The nucleoside diphosphate kinase from mimivirus: a peculiar affinity for deoxypyrimidine nucleotides

The first viral Nucleoside Diphosphate Kinase was recently identified in the giant double-stranded DNA virus Acanthamoeba polyphag a Mimivirus (ApM). Here we report its expression and detailed biochemical characterization. NDK_apm exhibits unique features such as a shorter Kpn-loop, a structural motif previously reported to be part of the active site and involved in oligomer formation. Enzymatic activity measurements on the recombinant NDK_apm revealed its preferential affinity for deoxypyrimidine nucleotides. This property might represent an adaptation of NDK_apm to the production of the limiting TTP deoxynucleotide required for the replication of the large A+T rich (72%) viral genome. The NDK_apm might also assume a role in dUTP detoxification to compensate for the surprising absence of Mimivirus dUTPase (deoxyuridine triphosphate pyrophosphatase) an important enzyme conserved in most viruses. Although the phylogenetic analysis of NDK sequences sampled through organisms from the three domains of life is only partially informative, it favors an ancestral origin for NDK_apm over a recent acquisition from a eukaryotic organism by horizontal gene transfer.     

Molecular Diversity of New Thermococcales Isolates from a Single Area of Hydrothermal Deep-Sea Vents as Revealed by Randomly Amplified Polymorphic DNA Fingerprinting and 16S rRNA Gene Sequence Analysis

Members of the Thermococcales are anaerobic Archaea belonging to the kingdom Euryarchaea that are studied in many laboratories as model organisms for hyperthermophiles. We describe here a molecular analysis of 86 new Thermococcales isolates collected from six different chimneys of a single hydrothermal field located in the 13°N 104°W segment of the East Pacific ridge at a depth of 2,330 m. These isolates were sorted by randomly amplified polymorphic DNA (RAPD) fingerprinting into nine groups, and nine unique RAPD profiles were obtained. One RAPD group corresponds to new isolates of Thermococcus hydrothermalis, whereas all other groups and isolates with unique profiles are different from the 22 reference strains included in this study. Analysis of 16S rRNA gene sequences of representatives of each RAPD group and unique profiles showed that one group corresponds to Pyrococcus strains, whereas all the other isolates are Thermococcus strains. We estimated that our collection may contain at least 11 new species. These putative species, isolated from a single area of hydrothermal deep-sea vents, are dispersed in the 16S rRNA tree among the reference strains previously isolated from diverse hot environments (terrestrial, shallow water, hydrothermal vents) located around the world, suggesting that there is a high degree of dispersal of Thermococcales. About one-half of our isolates contain extrachromosomal elements that could be used to search for novel replication proteins and to develop genetic tools for hyperthermophiles.     

Construction of Chromosome Segment Substitution Lines in Peanut (Arachis hypogaea L.) Using a Wild Synthetic and QTL Mapping for Plant Morphology

Chromosome segment substitution lines (CSSLs) are powerful QTL mapping populations that have been used to elucidate the molecular basis of interesting traits of wild species. Cultivated peanut is an allotetraploid with limited genetic diversity. Capturing the genetic diversity from peanut wild relatives is an important objective in many peanut breeding programs. In this study, we used a marker-assisted backcrossing strategy to produce a population of 122 CSSLs from the cross between the wild synthetic allotetraploid (A. ipaënsis×A. duranensis)^4x and the cultivated Fleur11 variety. The 122 CSSLs offered a broad coverage of the peanut genome, with target wild chromosome segments averaging 39.2 cM in length. As a demonstration of the utility of these lines, four traits were evaluated in a subset of 80 CSSLs. A total of 28 lines showed significant differences from Fleur11. The line×trait significant associations were assigned to 42 QTLs: 14 for plant growth habit, 15 for height of the main stem, 12 for plant spread and one for flower color. Among the 42 QTLs, 37 were assigned to genomic regions and three QTL positions were considered putative. One important finding arising from this QTL analysis is that peanut growth habit is a complex trait that is governed by several QTLs with different effects. The CSSL population developed in this study has proved efficient for deciphering the molecular basis of trait variations and will be useful to the peanut scientific community for future QTL mapping studies.     

Insights into the binding and activation sites of the receptors for cholecystokinin and gastrin

CCK receptors represent potential targets in a number of diseases. Knowledge of CCK receptor binding sites is a prerequisite for the understanding of the molecular basis for their ligand recognition, partial agonism, ligand-induced trafficking of signalling. In the current paper, we report studies from our laboratory and others which have provided new data on the molecular basis of the pharmacology and functioning of CCK1 and CCK2 receptors. It has been shown that: 1) homologous regions of the two receptors are involved in the binding site of CCK, however, positioning of CCK slightly differs in agreement with distinct pharmacophores of CCK toward the two receptors and receptor sequence variations; 2) Binding sites of most of non-peptide agonists/ antagonist are buried in the pocket formed by transmembrane helices and overlap that of CCK; Aromatic amino acids within and near the binding site, especially in helix VI, are involved in receptor activation; 4) Like for other members of family A of G-protein coupled receptors, residues of the binding sites as well as of conserved motifs such as E/DRY, NPXXY are crucial for receptor activation.     

Genome Scale Evolution of Myxoma Virus Reveals Host-Pathogen Adaptation and Rapid Geographic Spread

The evolutionary interplay between myxoma virus (MYXV) and the European rabbit (Oryctolagus cuniculus) following release of the virus in Australia in 1950 as a biological control is a classic example of host-pathogen coevolution. We present a detailed genomic and phylogeographic analysis of 30 strains of MYXV, including the Australian progenitor strain Standard Laboratory Strain (SLS), 24 Australian viruses isolated from 1951 to 1999, and three isolates from the early radiation in Britain from 1954 and 1955. We show that in Australia MYXV has spread rapidly on a spatial scale, with multiple lineages cocirculating within individual localities, and that both highly virulent and attenuated viruses were still present in the field through the 1990s. In addition, the detection of closely related virus lineages at sites 1,000 km apart suggests that MYXV moves freely in geographic space, with mosquitoes, fleas, and rabbit migration all providing means of transport. Strikingly, despite multiple introductions, all modern viruses appear to be ultimately derived from the original introductions of SLS. The rapidity of MYXV evolution was also apparent at the genomic scale, with gene duplications documented in a number of viruses. Duplication of potential virulence genes may be important in increasing the expression of virulence proteins and provides the basis for the evolution of novel functions. Mutations leading to loss of open reading frames were surprisingly frequent and in some cases may explain attenuation, but no common mutations that correlated with virulence or attenuation were identified.     

“Hidden” dUTPase Sequence in Human Immunodeficiency Virus Type 1 gp120

A coding region homologous to the sequence for essential eukaryotic enzyme dUTPase has been identified in different genomic regions of several viral lineages. Unlike the nonprimate lentiviruses (caprine arthritis- encephalitis virus, equine infectious anemia virus, feline immunodeficiency virus, and visna virus), where dUTPase is integrated into the pol coding region, this enzyme has never been demonstrated to be present in the primate lentivirus genomes (human immunodeficiency virus type 1 [HIV-1], HIV-2, or the related simian immunodeficiency virus). A novel approach allowed us to identify a weak but significant sequence similarity between HIV-1 gp120 and the human dUTPase. This finding was then extended to all of the primate lentivirus lineages. Together with the recently reported fragmentary structural similarity between the V3 loop region and the Escherichia coli dUTPase (P. D. Kwong, R. Wyatt, J. Robinson, R. W. Sweet, J. Sodroski, and W. A. Hendrickson, Nature 393:648–659, 1998), our results strongly suggest that an ancestral dUTPase gene has evolved into the present primate lentivirus CD4 and cytokine receptor interacting region of gp120.     

Bovine-like coronaviruses isolated from four species of captive wild ruminants are homologous to bovine coronaviruses, based on complete genomic sequences

We sequenced and analyzed the full-length genomes of four coronaviruses (CoVs), each from a distinct wild-ruminant species in Ohio: sambar deer (Cervus unicolor), a waterbuck (Kobus ellipsiprymnus), a sable antelope (Hippotragus niger), and a white-tailed deer (Odocoileus virginianus). The fecal samples from the sambar deer, the waterbuck, and the white-tailed deer were collected during winter dysentery outbreaks and sporadic diarrhea cases in 1993 and 1994 (H. Tsunemitsu, Z. R. el-Kanawati, D. R. Smith, H. H. Reed, and L. J. Saif, J. Clin. Microbiol. 33:3264-3269, 1995). A fecal sample from a sable antelope was collected in 2003 from an Ohio wild-animal habitat during the same outbreak when a bovine-like CoV from a giraffe (GiCoV) was isolated (M. Hasoksuz, K. Alekseev, A. Vlasova, X. Zhang, D. Spiro, R. Halpin, S. Wang, E. Ghedin, and L. J. Saif, J. Virol. 81:4981-4990, 2007). For two of the CoVs (sambar deer and waterbuck), complete genomes from both the cell culture-adapted and gnotobiotic-calf-passaged strains were also sequenced and analyzed. Phylogenetically, wild-ruminant CoVs belong to group 2a CoVs, with the closest relatedness to recent bovine CoV (BCoV) strains. High nucleotide identities (99.4 to 99.6%) among the wild-ruminant strains and recent BCoV strains (BCoV-LUN and BCoV-ENT, isolated in 1998) further confirm the close relatedness. Comparative genetic analysis of CoVs of captive wild ruminants with BCoV strains suggests that no specific genomic markers are present that allow discrimination between the bovine strains and bovine-like CoVs from captive wild ruminants; furthermore, no specific genetic markers were identified that defined cell cultured or calf-passaged strains or the host origin of strains. The results of this study confirm prior reports of biologic and antigenic similarities between bovine and wild-ruminant CoVs and suggest that cattle may be reservoirs for CoVs that infect captive wild ruminants or vice versa and that these CoVs may represent host range variants of an ancestral CoV.     

Telomere Length Correlations among Somatic Tissues in Adult Zebra Finches

Telomeres are repetitive non coding DNA sequences located at the end of eukaryotic chromosomes, which maintain the integrity of the genome by hiding the chromosome ends from being recognised as double stranded breaks. Telomeres are emerging as biomarkers for ageing and survival, and are susceptible to reflect different individual life history trajectories. In particular, the telomere length with which one starts in life has been shown to be linked with individual life-long survival, suggesting that telomere dynamics can be a proxy for individual fitness and thereby be implicated in evolutionary trade-offs. As a consequence, an increasing number of studies were conducted on telomeres in the fields of ecology and evolutionary biology, in which telomere length was almost exclusively measured from blood samples. However, not only do the number of repeats of the telomeric sequences vary among species, but also within species with great inter-individual telomere lengths variability with age, tissues, and chromosomes. This raises the issue of the exact biological meaning of telomere measurement in blood cells and stimulated the study of the correlation of telomere lengths among tissues over age. By measuring telomere length in adult zebra finches (Taeniopygia guttata) in different somatic tissues displaying variable cell turnovers (bone marrow, brain, spleen, pectoral muscle, heart, liver and in red blood cells), we checked that the measure of telomere length in red blood cells is related to telomere lengths in the other tissues. Here we show significant relationships between the telomere lengths of red blood cells and several somatic tissues at adulthood. As red blood cells are easily accessible and suitable for the longitudinal monitoring of the individual rate of telomere loss, our study confirms that telomere length measured in red blood cells could serve as a surrogate for telomere length in the whole avian organism.