Phylogenetic, genomic organization and expression analysis of hydrophobin genes in the ectomycorrhizal basidiomycete Laccaria bicolor

Hydrophobins are morphogenetic, small secreted hydrophobic fungal proteins produced in response to changing development and environmental conditions. These proteins are important in the interaction between certain fungi and their hosts. In mutualistic ectomycorrhizal fungi several hydrophobins form a subclass of mycorrhizal-induced small secreted proteins that are likely to be critical in the formation of the symbiotic interface with host root cells. In this study, two genomes of the ectomycorrhizal basidiomycete Laccaria bicolor strains S238N-H82 (from North America) and 81306 (from Europe) were surveyed to construct a comprehensive genome-wide inventory of hydrophobins and to explore their characteristics and roles during host colonization. The S238N-H82 L. bicolor hydrophobin gene family is composed of 12 genes while the 81306 strain encodes nine hydrophobins, all corresponding to class I hydrophobins. The three extra hydrophobin genes encoded by the S238N-H82 genome likely arose via gene duplication and are bordered by transposon rich regions. Expression profiles of the hydrophobin genes of L. bicolor varied greatly depending on life stage (e.g. free living mycelium vs. root colonization) and on the host root environment. We conclude from this study that the complex diversity and range of expression profiles of the Laccaria hydrophobin multi-gene family have likely been a selective advantage for this mutualist in colonizing a wide range of host plants.     

Centrosome positioning in polarized cells: Common themes and variations

The centrosome position is tightly regulated during the cell cycle and during differentiated cellular functions. Because centrosome organizes the microtubule network to coordinate both intracellular organization and cell signaling, centrosome positioning is crucial to determine either the axis of cell division, the direction of cell migration or the polarized immune response of lymphocytes. Since alteration of centrosome positioning seems to promote cell transformation and tumor spreading, the molecular mechanisms controlling centrosome movement in response to extracellular and intracellular cues are under intense investigation. Evolutionary conserved pathways involving polarity proteins and cytoskeletal rearrangements are emerging as common regulators of centrosome positioning in a wide variety of cellular contexts.     

On the Relationship Between Clonal Traits and Small-Scale Spatial Patterns of Three Dominant Grasses and its Consequences on Community Diversity

In a secondary successional community, we focused on the role of local dispersion mediated by clonal growth in the density and spatial patterning of tillers of three dominant grass species (Elymus repens, Brachypodium pinnatum and Calamagrostis epigejos) on the plant neighbourhood scale. We also asked whether the spatial pattern/density of their tillers were linked to the local diversity structure. In ten 75 cm?×?75 cm quadrats for each of the three species, we quantified i) the clonal morphology patterns from measuring spacer length, branching rate and the number of clumping tillers per module, ii) the spatial patterns and density of tillers in grids at four different resolutions (cell sizes varied between 1 cm?×?1 cm; 2.5 cm?×?2.5 cm; 5 cm?×?5 cm and 7.5 cm?×?7.5 cm), and iii) local species richness and local dominance based on botanical relevés. Then, we explored the relationships between iv) the clonal architecture pattern and the density/spatial pattern of tillers and v) the density/spatial pattern of tillers and local diversity variables, through regression analyses. Aggregation intensity on the smallest scales and tiller density were negatively linked to spacer length and positively linked to branching rate and number of clumping tillers. Species richness and dominance in quadrats were negatively and positively linked to tiller density, respectively. Dominance was positively linked to aggregation intensity on a 1-cm scale. This study emphasized and quantified the importance of clonal growth in the intensity and quality of grass tiller patterning in space on the plant neighbourhood scale. Our approach allowed the accurate positioning of species, or even clones on the phalanx-guerrilla continuum. This should help us to understand how dominant grass species affect the dynamics of stand communities.     

Isolation and characterization of Tn-Dha1, a transposon containing the tetrachloroethene reductive dehalogenase of Desulfitobacterium hafniense strain TCE1

A new 9.9 kb catabolic transposon, Tn-Dha1, containing the gene responsible for tetrachloroethene (PCE) reductive dechlorination activity, was isolated from Desulfitobacterium hafniense strain TCE1. Two fully identical copies of the insertion sequence ISDha1, a new member of the IS256 family, surround the gene cluster pceABCT, a truncated gene for another transposase and a short open reading frame with homology to a member of the twin-arginine transport system (tatA). Evidence was obtained by Southern blot for an alternative form of the transposon element as a circular molecule containing only one copy of ISDha1. This latter structure most probably represents a dead-end product of the transposition of Tn-Dha1. Strong indications for the transposition activity of ISDha1 were given by polymerase chain reaction (PCR) amplification and sequencing of the intervening sequence located between both inverted repeats (IR) of ISDha1 (IR junction). A stable genomic ISDha1 tandem was excluded by quantitative real-time PCR. Promoter mapping of the pceA gene, encoding the reductive dehalogenase, revealed the presence of a strong promoter partially encoded in the right inverted repeat of ISDha1. A sequence comparison with pce gene clusters from Desulfitobacterium sp. strains PCE-S and Y51 and from Dehalobacter restrictus, all of which show 100% identity for the pceAB genes, indicated that both Desulfitobacterium strains seem to possess the same transposon structure, whereas only the pceABCT gene cluster is conserved in D. restrictus.     

Linking two immuno-suppressive molecules: indoleamine 2,3 dioxygenase can modify HLA-G cell-surface expression

Nonclassical human leukocyte antigen (HLA) class I molecule HLA-G and indoleamine 2,3 dioxygenase (INDO) in humans and mice, respectively, have been shown to play crucial immunosuppressive roles in fetal-maternal tolerance. HLA-G inhibits natural killer and T cell function by high-affinity interaction with inhibitory receptors, and INDO acts by depleting the surrounding microenvironment of the essential amino acid tryptophan, thus inhibiting T cell proliferation. We investigated whether HLA-G expression and INDO function were linked. Working with antigen-presenting cell (APC) lines and monocytes, we found that functional inhibition of INDO by 1-methyl-tryptophan induced cell surface expression of HLA-G1 by HLA-G1-negative APCs that were originally cell-surface negative, and that in reverse, the functional boost of INDO by high concentrations of tryptophan induced a complete loss of HLA-G1 cell surface expression by APCs that were originally cell-surface HLA-G1-positive. This mechanism was shown to be posttranslational because HLA-G protein cell contents remained unaffected by the treatments used. Furthermore, HLA-G cell surface expression regulation by INDO seems to relate to INDO function, but not to tryptophan catabolism itself. Potential implications in fetal-maternal tolerance are discussed.     

Morphological, chemical and genetic differentiation of two subspecies of Cistus creticus L. (C. creticus subsp. eriocephalus and C. creticus subsp. corsicus)

Cistus creticus L., an aromatic species from the Mediterranean area, contains various diterpenes bearing the labdane skeleton. The production of essential oil from this species has potential economic value, but so far, it has not been optimized. In order to contribute to a better knowledge of this species and to its differentiation, the morphological characters, volatile chemical composition and genetic data of two subspecies (C. creticus subsp. eriocephalus and C. creticus subsp. corsicus) were investigated. The leaf trichomes were studied using scanning electron microscopy. The chemical composition of Corsican essential oil (C. creticus subsp. corsicus) has been reported using GC, GC/MS and ¹³C NMR; the main constituents were oxygenated labdane diterpenes (33.9%) such as 13-epi-manoyl oxide (18.5%). Using plant material (54 samples) collected from 18 geographically distinct areas of the islands of Corsica and Sardinia, the basis of variation in the headspace solid-phase microextraction volatile fraction and an inter-simple sequence repeat genetic analysis were also examined. It was shown that the two subspecies of C. creticus differed in morphology, essential oil production, volatile fraction composition and genetic data.     

Amino acid residues critical for RNA-binding in the N-terminal domain of the nucleocapsid protein are essential determinants for the infectivity of coronavirus in cultured cells

The N-terminal domain of the coronavirus nucleocapsid (N) protein adopts a fold resembling a right hand with a flexible, positively charged β-hairpin and a hydrophobic palm. This domain was shown to interact with the genomic RNA for coronavirus infectious bronchitis virus (IBV) and severe acute respiratory syndrome coronavirus (SARS-CoV). Based on its 3D structure, we used site-directed mutagenesis to identify residues essential for the RNA-binding activity of the IBV N protein and viral infectivity. Alanine substitution of either Arg-76 or Tyr-94 in the N-terminal domain of IBV N protein led to a significant decrease in its RNA-binding activity and a total loss of the infectivity of the viral RNA to Vero cells. In contrast, mutation of amino acid Gln-74 to an alanine, which does not affect the binding activity of the N-terminal domain, showed minimal, if any, detrimental effect on the infectivity of IBV. This study thus identifies residues critical for RNA binding on the nucleocapsid surface, and presents biochemical and genetic evidence that directly links the RNA binding capacity of the coronavirus N protein to the viral infectivity in cultured cells. This information would be useful in development of preventive and treatment approaches against coronavirus infection.     

Inactivation of poliovirus 1 and F-specific RNA phages and degradation of their genomes by UV irradiation at 254 nanometers

Several models (animal caliciviruses, poliovirus 1 [PV1], and F-specific RNA bacteriophages) are usually used to predict inactivation of nonculturable viruses. For the same UV fluence, viral inactivation observed in the literature varies from 0 to 5 logs according to the models and the methods (infectivity versus molecular biology). The lack of knowledge concerning the mechanisms of inactivation due to UV prevents us from selecting the best model. In this context, determining if viral genome degradation may explain the loss of infectivity under UV radiation becomes essential. Thus, four virus models (PV1 and three F-specific RNA phages: MS2, GA, and Qbeta) were exposed to UV radiation from 0 to 150 mJ.cm-2. PV1 is the least-resistant virus, while MS2 and GA phages are the most resistant, with phage Qbeta having an intermediate sensitivity; respectively, 6-log, 2.3-log, 2.5-log, and 4-log decreases for 50 mJ.cm-2. In parallel, analysis of RNA degradation demonstrated that this phenomenon depends on the fragment size for PV1 as well as for MS2. Long fragments (above 2,000 bases) for PV1 and MS2 fell rapidly to the background level (>1.3-log decrease) for 20 mJ.cm-2 and 60 mJ.cm-2, respectively. Nevertheless, the size of the viral RNA is not the only factor affecting UV-induced RNA degradation, since viral RNA was more rapidly degraded in PV1 than in the MS2 phage with a similar size. Finally, extrapolation of inactivation and UV-induced RNA degradation kinetics highlights that genome degradation could fully explain UV-induced viral inactivation.     

Trogocytosis-based generation of suppressive NK cells

Trogocytosis is a fast uptake of membranes and associated molecules from one cell by another. Trogocytosis between natural killer (NK) cells and tumors is already described, but the functional relevance of NK-tumor targets material exchange is unclear. We investigated whether the immunosuppressive molecule HLA-G that is commonly expressed by tumors in vivo and known to block NK cytolytic function, could be transferred from tumor cells to NK cells, and if this transfer had functional consequences. We show that activated NK cells acquire HLA-G1 from tumor cells, and that upon this acquisition, NK cells stop proliferating, are no longer cytotoxic, and behave as suppressor cells. Such cells can inhibit other NK cells' cytotoxic function and protect NK-sensitive tumor cells from cytolysis. These data are the first demonstration that trogocytosis of HLA-G1 can be a major mechanism of immune escape that acts through effector cells made to act as suppressor cells locally, temporarily, but efficiently. The broader consequences of membrane sharing between immune and non-immune cells on the function of effectors and the outcome of immune responses are discussed.