Rab11 Regulates Trafficking of Trans-sialidase to the Plasma Membrane through the Contractile Vacuole Complex of Trypanosoma cruzi

Trypanosoma cruzi is the etiologic agent of Chagas disease. Although this is not a free-living organism it has conserved a contractile vacuole complex (CVC) to regulate its osmolarity. This obligate intracellular pathogen is, in addition, dependent on surface proteins to invade its hosts. Here we used a combination of genetic and biochemical approaches to delineate the contribution of the CVC to the traffic of glycosylphosphatidylinositol (GPI)-anchored proteins to the plasma membrane of the parasite and promote host invasion. While T. cruzi Rab11 (GFP-TcRab11) localized to the CVC, a dominant negative (DN) mutant tagged with GFP (GFP-TcRab11DN) localized to the cytosol, and epimastigotes expressing this mutant were less responsive to hyposmotic and hyperosmotic stress. Mutant parasites were still able to differentiate into metacyclic forms and infect host cells. GPI-anchored trans-sialidase (TcTS), mucins of the 60–200 KDa family, and trypomastigote small surface antigen (TcTSSA II) co-localized with GFP-TcRab11 to the CVC during transformation of intracellular amastigotes into trypomastigotes. Mucins of the gp35/50 family also co-localized with the CVC during metacyclogenesis. Parasites expressing GFP-TcRab11DN prevented TcTS, but not other membrane proteins, from reaching the plasma membrane, and were less infective as compared to wild type cells. Incubation of these mutants in the presence of exogenous recombinant active, but not inactive, TcTS, and a sialic acid donor, before infecting host cells, partially rescued infectivity of trypomastigotes. Taking together these results reveal roles of TcRab11 in osmoregulation and trafficking of trans-sialidase to the plasma membrane, the role of trans-sialidase in promoting infection, and a novel unconventional mechanism of GPI-anchored protein secretion.     

Intraspecific phenotypic variability of plant functional traits in contrasting mountain grasslands habitats

Empirical studies that link plants intraspecific variation to environmental conditions are almost lacking, despite their relevance in understanding mechanisms of plant adaptation, in predicting the outcome of environmental change and in conservation. Here, we investigate intraspecific trait variation of four grassland species along with abiotic environmental variation at high spatial resolution (n = 30 samples per species trait and environmental factor per site) in two contrasting grassland habitats in Central Apennines (Italy). We test for phenotypic adaptation between habitats, intraspecific trait-environment relationships within habitats, and the extent of trait and environmental variation. We considered whole plant, clonal, leaf, and seed traits. Differences between habitats were tested using ANOVA and ANCOVA. Trait-environment relationships were assessed using multiple regression models and hierarchical variance partitioning. The extent of variation was calculated using the coefficient of variation. Significant intraspecific differences in trait attributes between the contrasting habitats indicate phenotypic adaptation to in situ environmental conditions. Within habitats, light, soil temperature, and the availability of nitrate, ammonium, magnesium and potassium were the most important factors driving intraspecific trait-environment relationships. Leaf traits and height growth show lower variability than environment being probably more regulated by plants than clonal traits which show much higher variability. We show the adaptive significance of key plant traits leading to intraspecific adaptation of strategies providing insights for conservation of extant grassland communities. We argue that protecting habitats with considerable medium- and small-scale environmental heterogeneity is important to maintain large intraspecific variability within local populations that finally can buffer against uncertainty of future climate and land use scenarios.     

A superfamily of Trypanosoma cruzi surface antigens

Several genes o f Trypanosoma cruzi encode surface antigens that include an amino acid motif that is conserved among bacterial neurominidases. Oscar Campetella, Daniel Sdnchez, Juan Jose Cazzulo and Alberto Carlos Frasch here suggest grouping these gene families in a superfamily.     

Contrasting patterns in leaf traits of Mediterranean shrub communities along an elevation gradient: measurements matter

Plant Ecology - We assessed the changes in community-weighted mean (CWM) and variability of specific leaf area (SLA) and leaf area (LA) of different Mediterranean shrub communities along an...     

Patterns of Clonal Growth Modes Along a Chronosequence of Post-Coppice Forest Regeneration in Beech Forests of Central Italy

Forest coppicing leads to changes in composition of the herbaceous understory through soil disturbance and alteration of the light regime. While the role of seed dispersal traits at the start of succession after coppicing has been extensively studied, the role of persistence traits such as clonal growth and bud banks is not yet sufficiently understood. To gain better understanding of this role, we studied the patterns of clonal growth organs and related clonal traits of species in a series of coppiced beech forests of the Central Apennines (Marches region, Italy) in various stages of recovery after the last coppicing event. We conducted stratified random sampling and established a chronosequence of recovery stages based on stand age (reflecting the number of years since the last coppicing). The beech stands were classified into three age groups (Post-logged, Recovering, and Old-coppice stands) according to the characteristic stages of beech coppice dynamics. Clonal growth organs and the corresponding clonal traits of plants in the forest understory vegetation were assessed with the help of a CLO-PLA1 database. We found no significant change in the proportion of clonal species along the studied chronosequence. In contrast, most of the traits and about the half of the clonal growth organs showed correlation with stand age or preference for a certain habitat (i.e., stage of regeneration). Clonal and bud bank traits proved to play an important role in the persistence of species subjected to forest coppicing cycles in the studied area.     

Unravelling mechanisms of short-term vegetation dynamics in complex coppice forest systems

The silvicultural management of coppicing has been very common in deciduous forests in many European countries. After decades of decline of this practice, socio-economic changes might induce a revival valuing the biomass as a resource. New insights in the ecological processes that regulate plant diversity are relevant for a sustainable forest management. While studies on long-term changes are available, the short-term dynamics of the coppice forest understorey has not yet been explored. In this context, it is interesting to evaluate the species compositional changes, including the processes of species turnover and species impoverishment (nestedness) and to investigate the role of plant functional traits. For this purpose, we resampled a chronosequence of complex coppice beech forests of the Central Apennines (Italy) monitoring the short-time species dynamics of five years (i.e. from 2006 to 2011) in three age classes, i.e. post-logged, recovering and old coppice stands (0–16, 17–31 and > 32 years, respectively). In contrast to our expectation, declining species richness appeared only in the recovering stands, while the landscape scale (between-stand) heterogeneity, except for post-logged and recovering stands in 2011, did not change over five years. Significant temporal nestedness was found in each stage of succession. However, the rate of species turnover and species impoverishment do not significantly differ among the three age classes, indicating their constant importance along the forest regeneration after disturbance. Only in the early stage of forest regeneration after coppicing, species compositional changes are reflected by functional changes with surviving understorey species having clonal regeneration traits. Our results suggest an overall landscape-scale stability (and sustainability) of this coppice forest system. We conclude with management indications, highlighting the importance of maintaining the traditional local approach (coppicing with standards in small 0.5–1.0 ha sized management units with a ca 30-year rotation cycle) where active coppice parcels are interspersed by abandoned stands.     

Effects of extreme drought on specific leaf area of grassland species: A meta‐analysis of experimental studies in temperate and sub‐Mediterranean systems

Here, we conducted a meta‐analysis of experimental drought manipulation studies using rainout shelters in five sites of natural grassland ecosystems of Europe. The single studies assess the effects of extreme drought on the intraspecific variation of the specific leaf area (SLA), a proxy of plant growth. We evaluate and compare the effect size of the SLA response for the functional groups of forbs and grasses in temperate and sub‐Mediterranean systems. We hypothesized that the functional groups of grasses and forbs from temperate grassland systems have different strategies in short‐term drought response, measured as adjustment of SLA, with SLA‐reduction in grasses and SLA‐maintenance in forbs. Second, we hypothesized that grasses and forbs from sub‐Mediterranean systems do not differ in their drought response as both groups maintain their SLA. We found a significant decrease of SLA in grasses of the temperate systems in response to drought while SLA of forbs showed no significant response. Lower SLA is associated with enhanced water‐use efficiency under water stress and thus can be seen as a strategy of phenotypic adjustment. By contrast, in the sub‐Mediterranean systems, grasses significantly increased their SLA in the drought treatment. This result points towards a better growth performance of these grasses, which is most likely related to their strategy to allocate resources to belowground parts. The observed SLA reduction of forbs is most likely a direct drought response given that competitive effect of grasses is unlikely due to the scanty vegetation cover. We point out that phenotypic adjustment is an important driver of short‐term functional plant response to climatic extremes such as drought. Differential reactions of functional groups have to be interpreted against the background of the group's evolutionary configuration that can differ between climatic zones.     

Structural basis of sialyltransferase activity in trypanosomal sialidases

The intracellular parasite Trypanosoma cruzi, the etiological agent of Chagas disease, sheds a developmentally regulated surface trans-sialidase, which is involved in key aspects of parasite-host cell interactions. Although it shares a common active site architecture with bacterial neuraminidases, the T.cruzi enzyme behaves as a highly efficient sialyltransferase. Here we report the crystal structure of the closely related Trypanosoma rangeli sialidase and its complex with inhibitor. The enzyme folds into two distinct domains: a catalytic beta-propeller fold tightly associated with a lectin-like domain. Comparison with the modeled structure of T.cruzi trans-sialidase and mutagenesis experiments allowed the identification of amino acid substitutions within the active site cleft that modulate sialyltransferase activity and suggest the presence of a distinct binding site for the acceptor carbohydrate. The structures of the Trypanosoma enzymes illustrate how a glycosidase scaffold can achieve efficient glycosyltransferase activity and provide a framework for structure-based drug design.