Large-scale discovery and characterization of protein regulatory motifs in eukaryotes

The increasing ability to generate large-scale, quantitative proteomic data has brought with it the challenge of analyzing such data to discover the sequence elements that underlie systems-level protein behavior. Here we show that short, linear protein motifs can be efficiently recovered from proteome-scale datasets such as sub-cellular localization, molecular function, half-life, and protein abundance data using an information theoretic approach. Using this approach, we have identified many known protein motifs, such as phosphorylation sites and localization signals, and discovered a large number of candidate elements. We estimate that ~80% of these are novel predictions in that they do not match a known motif in both sequence and biological context, suggesting that post-translational regulation of protein behavior is still largely unexplored. These predicted motifs, many of which display preferential association with specific biological pathways and non-random positioning in the linear protein sequence, provide focused hypotheses for experimental validation.     

Distribution and biochemical properties of an M1-family aminopeptidase in Plasmodium falciparum indicate a role in vacuolar hemoglobin catabolism

Aminopeptidases catalyze N-terminal peptide bond hydrolysis and occupy many diverse roles across all domains of life. Here we present evidence that an M1-family aminopeptidase, PfA-M1, has been recruited to specialized roles in the human malaria parasite Plasmodium falciparum. PfA-M1 is abundant in two subcellular compartments in asexual intraerythrocytic parasites; that is, the food vacuole, where the catabolism of host hemoglobin takes place, and the nucleus. A unique N-terminal extension contributes to the observed dual targeting by providing a signal peptide and putative alternate translation initiation sites. PfA-M1 exists as two major isoforms, a nuclear 120-kDa species and a processed species consisting of a complex of 68- and 35-kDa fragments. PfA-M1 is both stable and active at the acidic pH of the food vacuole lumen. Determination of steady-state kinetic parameters for both aminoacyl-β-naphthylamide and unmodified dipeptide substrates over the pH range 5.0-8.5 reveals that k(cat) is relatively insensitive to pH, whereas K(m) increases at pH values below 6.5. At the pH of the food vacuole lumen (5.0-5.5), the catalytic efficiency of PfA-M1 remains high. Consistent with the kinetic data, the affinity of peptidic competitive inhibitors is diminished at acidic pH. Together, these results support a catalytic role for PfA-M1 in the food vacuole and indicate the importance of evaluating the potency of peptidic inhibitors at physiologically relevant pH values. They also suggest a second, distinct function for this enzyme in the parasite nucleus.     

Immune evasion by Helicobacter pylori is mediated by induction of macrophage arginase II

Helicobacter pylori infection persists for the life of the host due to the failure of the immune response to eradicate the bacterium. Determining how H. pylori escapes the immune response in its gastric niche is clinically important. We have demonstrated in vitro that macrophage NO production can kill H. pylori, but induction of macrophage arginase II (Arg2) inhibits inducible NO synthase (iNOS) translation, causes apoptosis, and restricts bacterial killing. Using a chronic H. pylori infection model, we determined whether Arg2 impairs host defense in vivo. In C57BL/6 mice, expression of Arg2, but not arginase I, was abundant and localized to gastric macrophages. Arg2(-/-) mice had increased histologic gastritis and decreased bacterial colonization compared with wild-type (WT) mice. Increased gastritis scores correlated with decreased colonization in individual Arg2(-/-) mice but not in WT mice. When mice infected with H. pylori were compared, Arg2(-/-) mice had more gastric macrophages, more of these cells were iNOS(+), and these cells expressed higher levels of iNOS protein, as determined by flow cytometry and immunofluorescence microscopy. There was enhanced nitrotyrosine staining in infected Arg2(-/-) versus WT mice, indicating increased NO generation. Infected Arg2(-/-) mice exhibited decreased macrophage apoptosis, as well as enhanced IFN-γ, IL-17a, and IL-12p40 expression, and reduced IL-10 levels consistent with a more vigorous Th1/Th17 response. These studies demonstrate that Arg2 contributes to the immune evasion of H. pylori by limiting macrophage iNOS protein expression and NO production, mediating macrophage apoptosis, and restraining proinflammatory cytokine responses.     

Effect of equated continuous and interval running programs on endurance performance and jump capacity

We evaluated the effect of 2 different interval and continuous training programs on the maximal aerobic speed (MAS), time limit at MAS (T(lim)), and on the countermovement jump (CMJ). Twenty-two physically active men were randomly distributed in an interval training group (ITG), continuous training group (CTG), and control group. The CTG and ITG performed 2 different training programs (65-70 and 90-100% of the MAS for CTG and ITG, respectively) that consisted of 3 sessions per week during a period of 8 weeks with an identical external workload (% MAS × duration in minutes). The MAS, the T(lim) and the CMJ were recorded before and after the running training programs. The data analysis showed a significant and similar improvement (p < 0.01) of the MAS for both the ITG (5.8%) and CTG (8.3%). The T(lim) and CMJ did not change significantly for either group after the training period. Our results indicate that 8 weeks of continuous or interval running programs with externally equated load led to similar improvements in the MAS without changing T(lim) and CMJ performance in moderately trained nonrunners.     

Pooled genome-wide analysis to identify novel risk loci for pediatric allergic asthma

Background

Genome-wide association studies of pooled DNA samples were shown to be a valuable tool to identify candidate SNPs associated to a phenotype. No such study was up to now applied to childhood allergic asthma, even if the very high complexity of asthma genetics is an appropriate field to explore the potential of pooled GWAS approach.

Methodology/Principal Findings

We performed a pooled GWAS and individual genotyping in 269 children with allergic respiratory diseases comparing allergic children with and without asthma. We used a modular approach to identify the most significant loci associated with asthma by combining silhouette statistics and physical distance method with cluster-adapted thresholding. We found 97% concordance between pooled GWAS and individual genotyping, with 36 out of 37 top-scoring SNPs significant at individual genotyping level. The most significant SNP is located inside the coding sequence of C5, an already identified asthma susceptibility gene, while the other loci regulate functions that are relevant to bronchial physiopathology, as immune- or inflammation-mediated mechanisms and airway smooth muscle contraction. Integration with gene expression data showed that almost half of the putative susceptibility genes are differentially expressed in experimental asthma mouse models.

Conclusion/Significance

Combined silhouette statistics and cluster-adapted physical distance threshold analysis of pooled GWAS data is an efficient method to identify candidate SNP associated to asthma development in an allergic pediatric population.     

Potential Suitability and Viability of Selected Biodiesel Crops in Australian Marginal Agricultural Lands Under Current and Future Climates

The potential environmental suitability and economic viability of growing two biodiesel crops in marginal regions of Australia were explored. Firstly, we used spatial analysis techniques to identify marginal agricultural regions suitable for growing pongam (Pongamia pinnata) and Indian mustard (Brassica juncea), and determined the base socioeconomic viability of investments for the production of biodiesel in the identified areas. Secondly, we used climate change projections (target years 2020 to 2070) from the Commonwealth Scientific, Industrial and Research Organization Mk3.0 global circulation model generated for two emission scenarios (A1B and A1FI) to determine the shift in potential areas for these crops. Under the climate change scenarios tested, the total area suitable for growing pongam between 2040 and 2070 is substantially different from the suitable area under current climate, indicating that long-term investments in this perennial tree crop may not be viable in all regions, especially in southern Australia. There is a greater variation in suitability projections for Indian mustard, although there is more flexibility for cropping options given that it is an annual crop. However, future economic viability is likely to depend on the ability to receive renewable energy certificates for both crops and, in the case of pongam, the certified emission reductions. Opportunities exist for sustainable pongam agroforestry to supply biodiesel to regional towns, cattle stations and mines in northern Australia.