Fgf15 is required for proper morphogenesis of the mouse cardiac outflow tract

Evidence in animal models indicates that signaling networks functioning in the developing pharyngeal arches regulate stereotyped processes critical for proper development of the aortic arch and cardiac outflow tract. Here, we describe the phenotype of mice lacking fibroblast growth factor 15 (Fgf15), which encodes a secreted signaling molecule expressed within the developing pharyngeal arches. Homozygous Fgf15 mutants present heart defects consistent with malalignment of the aorta and pulmonary trunk. These defects correlate with early morphological defects of the outflow tract due to aberrant behavior of the cardiac neural crest. We demonstrate that Fgf15 expression within the pharyngeal arches is unaltered by a loss of Tbx1, a key regulator of pharyngeal arch development implicated in DiGeorge syndrome. In addition, Fgf15 and Tbx1 do not interact genetically, suggesting that Fgf15 operates through a pathway independent of Tbx1. These studies reveal a novel role of Fgf15 during development of the cardiac outflow tract.     

Functional conservation and specialization among eukaryotic anti-silencing function 1 histone chaperones

Chromatin disassembly and reassembly, mediated by histone chaperones such as anti-silencing function 1 (Asf1), are likely to accompany all nuclear processes that occur on the DNA template. In order to gain insight into the functional conservation of Asf1 across eukaryotes, we have replaced the budding yeast Asf1 protein with Drosophila Asf1 (dAsf1) or either of the two human Asf1 (hAsf1a and hAsf1b) counterparts. We found that hAsf1b is best able to rescue the growth defect of Saccharomyces cerevisiae lacking Asf1. Moreover, dAsf1 and hAsf1b but not hAsf1a can replace the role of yeast Asf1 in protecting against replicational stress and activating the PHO5 gene, while only hAsf1a can replace the role of Asf1 in protecting against double-stranded-DNA-damaging agents. Furthermore, it appears that the interaction between Asf1 and the DNA damage checkpoint protein Rad53 is not required for Asf1's role in maintaining genomic integrity. In addition to indicating the functional conservation of the Asf1 proteins across species, these studies suggest distinct roles for the two human Asf1 proteins.     

Sorghum genome sequencing by methylation filtration

Sorghum bicolor is a close relative of maize and is a staple crop in Africa and much of the developing world because of its superior tolerance of arid growth conditions. We have generated sequence from the hypomethylated portion of the sorghum genome by applying methylation filtration (MF) technology. The evidence suggests that 96% of the genes have been sequence tagged, with an average coverage of 65% across their length. Remarkably, this level of gene discovery was accomplished after generating a raw coverage of less than 300 megabases of the 735-megabase genome. MF preferentially captures exons and introns, promoters, microRNAs, and simple sequence repeats, and minimizes interspersed repeats, thus providing a robust view of the functional parts of the genome. The sorghum MF sequence set is beneficial to research on sorghum and is also a powerful resource for comparative genomics among the grasses and across the entire plant kingdom. Thousands of hypothetical gene predictions in rice and Arabidopsis are supported by the sorghum dataset, and genomic similarities highlight evolutionarily conserved regions that will lead to a better understanding of rice and Arabidopsis.     

An inducible translocation strategy to rapidly activate and inhibit small GTPase signaling pathways

We made substantial advances in the implementation of a rapamycin-triggered heterodimerization strategy. Using molecular engineering of different targeting and enzymatic fusion constructs and a new rapamycin analog, Rho GTPases were directly activated or inactivated on a timescale of seconds, which was followed by pronounced cell morphological changes. As signaling processes often occur within minutes, such rapid perturbations provide a powerful tool to investigate the role, selectivity and timing of Rho GTPase-mediated signaling processes.     

Integrating HIV prevention and treatment: from slogans to impact

Background

Through major efforts to reduce costs and expand access to antiretroviral therapy worldwide, widespread delivery of effective treatment to people living with HIV/AIDS is now conceivable even in severely resource-constrained settings. However, the potential epidemiologic impact of treatment in the context of a broader strategy for HIV/AIDS control has not yet been examined. In this paper, we quantify the opportunities and potential risks of large-scale treatment roll-out.

Methods and Findings

We used an epidemiologic model of HIV/AIDS, calibrated to sub-Saharan Africa, to investigate a range of possible positive and negative health outcomes under alternative scenarios that reflect varying implementation of prevention and treatment. In baseline projections, reflecting “business as usual,” the numbers of new infections and AIDS deaths are expected to continue rising. In two scenarios representing treatment-centered strategies, with different assumptions about the impact of treatment on transmissibility and behavior, the change in the total number of new infections through 2020 ranges from a 10% increase to a 6% reduction, while the number of AIDS deaths through 2020 declines by 9% to 13%. A prevention-centered strategy provides greater reductions in incidence (36%) and mortality reductions similar to those of the treatment-centered scenarios by 2020, but more modest mortality benefits over the next 5 to 10 years. If treatment enhances prevention in a combined response, the expected benefits are substantial—29 million averted infections (55%) and 10 million averted deaths (27%) through the year 2020. However, if a narrow focus on treatment scale-up leads to reduced effectiveness of prevention efforts, the benefits of a combined response are considerably smaller—9 million averted infections (17%) and 6 million averted deaths (16%). Combining treatment with effective prevention efforts could reduce the resource needs for treatment dramatically in the long term. In the various scenarios the numbers of people being treated in 2020 ranges from 9.2 million in a treatment-only scenario with mixed effects, to 4.2 million in a combined response scenario with positive treatment–prevention synergies.

Conclusions

These analyses demonstrate the importance of integrating expanded care activities with prevention activities if there are to be long-term reductions in the number of new HIV infections and significant declines in AIDS mortality. Treatment can enable more effective prevention, and prevention makes treatment affordable. Sustained progress in the global fight against HIV/AIDS will be attained only through a comprehensive response.     

Correlative GC-TOF-MS-based metabolite profiling and LC-MS-based protein profiling reveal time-related systemic regulation of metabolite–protein networks and improve pattern recognition for multiple biomarker selection

A novel approach is presented combining quantitative metabolite and protein data and multivariate statistics for the analysis of time-related regulatory effects of plant metabolism at a systems level. For the analysis of metabolites, gas chromatography coupled to a time-of-flight mass analyzer (GC-TOF-MS) was used. Proteins were identified and quantified using a novel procedure based on shotgun sequencing as described recently (Weckwerth etal., 2004b, Proteomics 4, 78–83). For comparison, leaves of Arabidopsis thaliana wild type plants and starchless mutant plants deficient in phosphoglucomutase activity (PGM) were sampled at intervals throughout the day/night cycle. Using principal and independent components analysis, each dataset (metabolites and proteins) displayed discrete characteristics. Compared to the analysis of only metabolites or only proteins, independent components analysis (ICA) of the integrated metabolite/protein dataset resulted in an improved ability to distinguish between WT and PGM plants (first independent component) and, in parallel, to see diurnal variations in both plants (second independent component). Interestingly, levels of photorespiratory intermediates such as glycerate and glycine best characterized phases of diurnal rhythm, and were not influenced by high sugar accumulation in PGM plants. In contrast to WT plants, PGM plants showed an inversely regulated cluster of N-rich amino acid metabolites and carbohydrates, indicating a shift in C/N partitioning. This observation corresponds to altered utilization of urea cycle intermediates in PGM plants suggesting enhanced protein degradation and carbon utilization due to growth inhibition. Among the proteins chloroplastidic GAPDH (At3g26650) was the best discriminator between WT and PGM plants in contrast to the cytosolic isoform (At1g13440) according to the primary effect of mutation located in the chloroplast. The described method is applicable to all kinds of biological systems and enables the unbiased identification of biomarkers embedded in correlative metabolite–protein networks.     

Animals as Sentinels of Human Environmental Health Hazards: An Evidence-Based Analysis

Despite recognition that animals could be serving as sentinels for environmental risks to human health, there are no evidence-based guidelines for the use of animal sentinel data in human health decision making. We performed a systematic review of the animal sentinel literature to assess the evidence linking such events to human health. A search of MEDLINE identified peer-reviewed original studies of animals as sentinels for either chemical or biological environmental hazards. A limited search of the CAB and AGRICOLA databases was also performed. We classified a random sample of 100 studies from the MEDLINE search according to species, hazard, and health outcome examined; study methods; and linkages to human health. Animal sentinel studies were difficult to locate in MEDLINE because of a lack of adequate key words for this concept. We found significant limitations in the study methods used to investigate animal sentinel events. Clear linkages to human health were frequently absent. Studies of sentinel events in animal populations hold potential for the recognition and control of human environmental health hazards, yet a number of barriers exist to using such data for evidence-based human health decisions. There is a need for greater data sharing and cooperative research between human and animal health professionals regarding environmental hazards and health outcomes in animal and human populations.     

A two-dimensional electrophoretic map of human mitochondrial proteins from immortalized lymphoblastoid cell lines: a prerequisite to study mitochondrial disorders in patients

Mitochondrial diseases may be caused by numerous mutations that alter proteins of the respiratory chain and of other metabolic pathways in the mitochondrium. For clinicians this disease group poses a considerable diagnostic challenge due to ambiguous genotype-phenotype relationships. Until now, only 30% of the mitochondriopathies can be diagnosed at the molecular level. We therefore need a new diagnostic tool that offers a wide view on the mitochondrial proteins. Here, we present a method to generate a high-resolution, large-gel two-dimensional gel electrophoretic (2-DE) map of a purified fraction of mitochondrial proteins from Epstein-Barr virus-immortalized lymphoblastoid cell line (LCL). LCLs can be easily obtained from patients and control subjects in a routine clinical setting. They often express the biochemical phenotype and can be cultured to high cell numbers, sufficient to gain enough purified material for 2-DE. In total we identified 166 mitochondrial proteins. Thirteen proteins were earlier not known to be of mitochondrial origin. Thirty-nine proteins were associated with human diseases ranging from respiratory chain enzyme deficiencies to disorders of beta-oxidation and amino acid metabolism. This 2-DE map is intended to be the first step to diagnose mitochondrial diseases at the proteomic level.     

An evaluation, comparison, and accurate benchmarking of several publicly available MS/MS search algorithms: sensitivity and specificity analysis

MS/MS and associated database search algorithms are essential proteomic tools for identifying peptides. Due to their widespread use, it is now time to perform a systematic analysis of the various algorithms currently in use. Using blood specimens used in the HUPO Plasma Proteome Project, we have evaluated five search algorithms with respect to their sensitivity and specificity, and have also accurately benchmarked them based on specified false-positive (FP) rates. Spectrum Mill and SEQUEST performed well in terms of sensitivity, but were inferior to MASCOT, X!Tandem, and Sonar in terms of specificity. Overall, MASCOT, a probabilistic search algorithm, correctly identified most peptides based on a specified FP rate. The rescoring algorithm, PeptideProphet, enhanced the overall performance of the SEQUEST algorithm, as well as provided predictable FP error rates. Ideally, score thresholds should be calculated for each peptide spectrum or minimally, derived from a reversed-sequence search as demonstrated in this study based on a validated data set. The availability of open-source search algorithms, such as X!Tandem, makes it feasible to further improve the validation process (manual or automatic) on the basis of "consensus scoring", i.e., the use of multiple (at least two) search algorithms to reduce the number of FPs. complement.