The HrpN effector of Erwinia amylovora, which is involved in type III translocation, contributes directly or indirectly to callose elicitation on apple leaves

Erwinia amylovora is responsible for fire blight of apple and pear trees. Its pathogenicity depends on a type III secretion system (T3SS) mediating the translocation of effectors into the plant cell. The DspA/E effector suppresses callose deposition on apple leaves. We found that E. amylovora and Pseudomonas syringae DC3000 tts mutants or peptide flg22 do not trigger callose deposition as strongly as the dspA/E mutant on apple leaves. This suggests that, on apple leaves, callose deposition is poorly elicited by pathogen-associated molecular patterns (PAMPs) such as flg22 or other PAMPs harbored by tts mutants and is mainly elicited by injected effectors or by the T3SS itself. Callose elicitation partly depends on HrpW because an hrpW-dspA/E mutant elicits lower callose deposition than a dspA/E mutant. Furthermore, an hrpN-dspA/E mutant does not trigger callose deposition, indicating that HrpN is required to trigger this plant defense reaction. We showed that HrpN plays a general role in the translocation process. Thus, the HrpN requirement for callose deposition may be explained by its role in translocation: HrpN could be involved in the translocation of other effectors inducing callose deposition. Furthermore, HrpN may also directly contribute to the elicitation process because we showed that purified HrpN induces callose deposition.     

Clonal Growth Strategies Along Flooding and Grazing Gradients in Atlantic Coastal Meadows

Specific composition and species clonal traits were characterized along combined flooding and grazing gradients to answer two questions. i) To what extent does the interaction of flooding and grazing influence the clonal characteristics of the vegetation? ii) Are the effects of both environmental factors independent or interactive? This study was carried out in a wet meadow along the Atlantic coast (France). Three plant communities (hygrophilous, meso-hygrophilous and mesophilous) were distinguished along a flooding gradient and five levels of grazing pressure were controlled through an experimental design (from no grazing to heavy grazing). We monitored species composition and retrieved, for each species, the type of clonal growth organs (CGOs) and clonal traits from the CLO-PLA3 database. We identified two syndromes of clonal traits: ??above-ground splitters?? and ??below-ground integrators??. Clonal traits played a key role in plant assembly in the studied meadows. The interaction of both environmental factors selected for particular syndromes of clonal traits; however, flooding had a stronger filtering effect than grazing. The hygrophilous community was dominated by above-ground splitters, whereas the meso-hygrophilous vegetation was dominated by below-ground integrators. In the mesophilous community, clonal composition was the most diverse and shared clonal traits with the vegetation of both the hygrophilous and meso-hygrophilous communities. Grazing impact on CGOs and clonal traits differed between plant communities, i.e., the effect of grazing was modulated by the flooding regime. This study confirmed that vegetation responses to grazing might depend on the pool of traits, primarily filtered by environmental factors such as flooding.     

Impact of selective grazing on plant production and quality through floristic contrasts and current-year defoliation in a wet grassland

Grazing impacts the structure and functional properties of vegetation through floristic changes (i.e., long-term effect) and current defoliation (i.e., short-term effect). The aim of this study was to assess the relative importance of these two grazing effects on productivity (ANPP) and plant quality (C/N ratio) among plant patches submitted to a variety of grazing intensity for several years. Long-term grazing effect was measured by comparing ANPP and C/N ratio among plant patches with contrasting floristic composition. Short-term impact of grazing was measured by comparing ANPP and C/N in plant patches, with and without defoliation. Floristic contrasts led to a lower ANPP in highly grazed patches than in lightly grazed ones. This result may be related to the increasing proportion of grazing-tolerant and grazing-avoiding species with increasing grazing intensity. Vegetation C/N contrasts were recorded among grazed patches but did not linearly relate to grazing intensity. Short-term effect of current-year defoliation on ANPP was limited as vegetation compensated for biomass removal. No evidence for grazing-enhancement of ANPP was found even at moderate grazing intensity. Long-term floristic changes with grazing thus appeared to be the main driving factor of variations in ANPP. In contrast, C/N ratio showed no general and consistent variation along the grazing gradient but varied consistently depending on the community investigated, thus suggesting an effect of the species pool available.     

Identification of proteins binding the native tubulin dimer

Microtubules play an essential role in eukaryotic cells, where they perform a wide variety of functions. In this paper, we describe the characterization of proteins associated to tubulin dimer in its native form, using affinity chromatography and mass spectrometry. We used an immunoaffinity column with coupled-monoclonal antibody directed against the alpha-tubulin C-terminus. Tubulin was first loaded onto the column, then interphase and mitotic cell lysates were chromatographed. Tubulin-binding proteins were eluted using a peptide mimicking the alpha-tubulin C-terminus. Elution fractions were analyzed by SDS-PAGE, and a total of 14 proteins were identified with high confidence by mass spectrometry. These proteins could be grouped in four classes: known tubulin-binding proteins, one microtubule-associated protein, heat shock proteins, and proteins that were not shown previously to bind tubulin dimer or microtubules.     

Endothelial stress induces the release of vitamin D-binding protein, a novel growth factor

Endothelial cells (EC) under stress release paracrine mediators that facilitate accumulation of vascular smooth muscle cells (VSCM) at sites of vascular injury. We found that medium conditioned by serum-starved EC increase proliferation and migration of VSCM in vitro. Fractionation of the conditioned medium followed by mass spectral analysis identified one bioactive component as vitamin D-binding protein (DBP). DBP induced both proliferation and migration of VSMC in vitro in association with increased phosphorylation of ERK 1/2. PD 98059, a biochemical inhibitor of ERK 1/2, abrogated these proliferative and migratory responses in VSMC. DBP is an important carrier for the vitamin-D sterols, 25-hydroxyvitamin-D, and 1alpha,25-dihydroxyvitamin-D. Both sterols inhibited the activity of DBP on VSMC, suggesting that vitamin D binding sites are important for initiating the activities of DBP on VSMC. Release of DBP at sites of endothelial injury represents a novel pathway favoring accumulation of VSMC at sites of vascular injury.     

Evidence of finely tuned expression of DNA polymerase beta in vivo using transgenic mice

DNA polymerase (Pol) is an error-prone repair DNA polymerase that has been shown to create genetic instability and tumorigenesis when overexpressed by only 2-fold in cells, suggesting that a rigorous regulation of its expression may be essential in vivo. To address this question, we have generated mice which express a transgene (Tg) bearing the Pol cDNA under the control of the ubiquitous promoter of the mouse H-2K gene from the major histocompatibility complex. These mice express the Tg only in thymus, an organ which normally contains the most abundant endogenous Pol mRNA and protein, supporting the idea of a tight regulation of Pol in vivo. Furthermore, we found no tumor incidence, suggesting that the single Pol overexpression event is not sufficient to initiate tumorigenesis in vivo.     

Evidence for genetic control of adult weight plasticity in the snail Helix aspersa

Phenotypic plasticity and canalization are important topics in quantitative genetics and evolution. Both concepts are related to environmental sensitivity. The latter can be modeled using a model with genetically structured environmental variance. This work reports the results of a genetic analysis of adult weight in the snail Helix aspersa. Several models of heterogeneous variance are fitted using a Bayesian, MCMC approach. Exploratory analyses using posterior predictive model checking and model comparisons based on the deviance information criterion favor a model postulating a genetically structured heterogeneous environmental variance. Our analysis provides a strong indication of a positive genetic correlation between additive genetic values affecting the mean and those affecting environmental variation of adult body weight. The possibility of manipulating environmental variance by selection is illustrated numerically using estimates of parameters derived from the snail data set.     

NARC-1/PCSK9 and its natural mutants: zymogen cleavage and effects on the low density lipoprotein (LDL) receptor and LDL cholesterol

The discovery of autosomal dominant hypercholesterolemic patients with mutations in the PCSK9 gene, encoding the proprotein convertase NARC-1, resulting in the missense mutations suggested a role in low density lipoprotein (LDL) metabolism. We show that the endoplasmic reticulum-localized proNARC-1 to NARC-1 zymogen conversion is Ca2+-independent and that within the zymogen autocatalytic processing site SSVFAQ [downward arrow]SIP Val at P4 and Pro at P3' are critical. The S127R and D374Y mutations result in approximately 50-60% and > or =98% decrease in zymogen processing, respectively. In contrast, the double [D374Y + N157K], F216L, and R218S natural mutants resulted in normal zymogen processing. The cell surface LDL receptor (LDLR) levels are reduced by 35% in lymphoblasts of S127R patients. The LDLR levels are also reduced in stable HepG2 cells overexpressing NARC-1 or its natural mutant S127R, and this reduction is abrogated in the presence of 5 mm ammonium chloride, suggesting that overexpression of NARC-1 increases the turnover rate of the LDLR. Adenoviral expression of wild type human NARC-1 in mice resulted in a maximal approximately 9-fold increase in circulating LDL cholesterol, while in LDLR-/- mice a delayed approximately 2-fold increase in LDL cholesterol was observed. In conclusion, NARC-1 seems to affect both the level of LDLR and that of circulating apoB-containing lipoproteins in an LDLR-dependent and -independent fashion.     

Population-based cervical screening with a 5-year interval in The Netherlands. Stabilization of the incidence of squamous cell carcinoma and its precursor lesions in the screened population

OBJECTIVE: To evaluate the outcome of population-based cervical screening at 5-year intervals. STUDY DESIGN: Results from the west region of the Netherlands (population 2 million) were used. The 1995-2000 round was compared with the first 2 years of the second (2001-2002). All results were prospectively collected in a central database. Positive cytologies and histoscores per 1,000 screened for preinvasive squamous cervical intraepithelial neoplasia (CIN) lesions and invasive squamous cell carcinoma were calculated. RESULTS: In the first round, 378,081 women were screened; in the second round, 100,561 women were screened. In both rounds the youngest screenees had the highest cytoscores. Cytoscores in the first round did not differ significantly from those in the second. The histoscore for CIN 1 and 2 was 1.42 per 1,000 in the first round and 1.18 per 1,000 (NS, P < .01) in the second. The histoscore for CIN 3 was 2.07 per 1,000 in the first round and 2.13 per 1,000 (NS, P < .01) in the second. Histoscores for invasive squamous cell carcinoma remained virtually the same (0.16 per 1,000 in the first, 0.14 per 1,000 in the second round). CONCLUSION: Population-based screening at 5-year intervals in the Netherlands may result in stabilization of positive cytology and of the incidence of CIN and (histologic) invasive squamous cell carcinoma. The program seems more cost effective than that of 2 decades ago, with a screening interval of 3 years.