Properties of purified actin depolymerizing factor from chick brain

Actin depolymerizing factor (ADF) from 19-day embryonic chick brains has been purified to greater than 98% homogeneity with a yield of 7.2 mg/100 g of brain. Quantitative immunoblotting with a monospecific antibody to ADF indicated that ADF comprises 0.3% of the total brain protein, resulting in an actual purification yield of about 20%. Brain ADF migrates as a single polypeptide of 19,000 kDa on SDS-containing polyacrylamide gels. The molecular weight of the native protein determined from sedimentation equilibrium in buffers containing from 50 to 200 mM KCl is 20,000. The secondary structure of ADF calculated from the circular dichroic spectrum consists of about 22% alpha-helix, 24% beta-sheet, and 18% beta-turn. ADF contains a blocked N-terminus, a single tryptophan residue located about one-third of the way from one end of the protein, and six cysteine residues (all in reduced form in the native protein). All six cysteine residues could be chemically modified with eosinylmaleimide under nondenaturing conditions; however, ADF activity was lost when more than one cysteine residue was modified. ADF microheterogeneity has been observed upon nonequilibrium pH gradient electrophoresis in polyacrylamide gels containing 9 M urea, the major isoform having a pI of congruent to 7.9-8.0. ADF can interact with either monomeric or filamentous actin to give a complex which can be isolated by gel filtration chromatography. Both major and minor isoforms of the ADF are found in the complex. Assembly-competent actin and active ADF can both be recovered from the complex by chromatography on ATP-saturated DEAE-cellulose.(ABSTRACT TRUNCATED AT 250 WORDS)     

Impact of artemisinin-based combination therapy and insecticide-treated nets on malaria burden in Zanzibar

Background

The Roll Back Malaria strategy recommends a combination of interventions for malaria control. Zanzibar implemented artemisinin-based combination therapy (ACT) for uncomplicated malaria in late 2003 and long-lasting insecticidal nets (LLINs) from early 2006. ACT is provided free of charge to all malaria patients, while LLINs are distributed free to children under age 5 y (“under five”) and pregnant women. We investigated temporal trends in Plasmodium falciparum prevalence and malaria-related health parameters following the implementation of these two malaria control interventions in Zanzibar.

Methods and Findings

Cross-sectional clinical and parasitological surveys in children under the age of 14 y were conducted in North A District in May 2003, 2005, and 2006. Survey data were analyzed in a logistic regression model and adjusted for complex sampling design and potential confounders. Records from all 13 public health facilities in North A District were analyzed for malaria-related outpatient visits and admissions. Mortality and demographic data were obtained from District Commissioner''s Office. P. falciparum prevalence decreased in children under five between 2003 and 2006; using 2003 as the reference year, odds ratios (ORs) and 95% confidence intervals (CIs) were, for 2005, 0.55 (0.28–1.08), and for 2006, 0.03 (0.00–0.27); p for trend < 0.001. Between 2002 and 2005 crude under-five, infant (under age 1 y), and child (aged 1–4 y) mortality decreased by 52%, 33%, and 71%, respectively. Similarly, malaria-related admissions, blood transfusions, and malaria-attributed mortality decreased significantly by 77%, 67% and 75%, respectively, between 2002 and 2005 in children under five. Climatic conditions favorable for malaria transmission persisted throughout the observational period.

Conclusions

Following deployment of ACT in Zanzibar 2003, malaria-associated morbidity and mortality decreased dramatically within two years. Additional distribution of LLINs in early 2006 resulted in a 10-fold reduction of malaria parasite prevalence. The results indicate that the Millennium Development Goals of reducing mortality in children under five and alleviating the burden of malaria are achievable in tropical Africa with high coverage of combined malaria control interventions.     

Assessment of genomic imprinting of SLC38A4, NNAT, NAP1L5, and H19 in cattle

Background

Sesame is an important oil crop in tropical and subtropical areas. Despite its nutritional value and historic and cultural importance, the research on sesame has been scarce, particularly as far as its genetic diversity is concerned. The aims of the present study were to clarify genetic relationships among 32 sesame accessions from the Venezuelan Germplasm Collection, which represents genotypes from five diversity centres (India, Africa, China-Korea-Japan, Central Asia and Western Asia), and to determine the association between geographical origin and genetic diversity using amplified fragment length polymorphism (AFLP).

Results

Large genetic variability was found within the germplasm collection. A total of 457 AFLP markers were recorded, 93 % of them being polymorphic. The Jaccard similarity coefficient ranged from 0.38 to 0.85 between pairs of accessions. The UPGMA dendrogram grouped 25 of 32 accessions in two robust clusters, but it has not revealed any association between genotype and geographical origin. Indian, African and Chinese-Korean-Japanese accessions were distributed throughout the dendrogram. A similar pattern was obtained using principal coordinates analysis. Genetic diversity studies considering five groups of accessions according to the geographic origin detected that only 20 % of the total diversity was due to diversity among groups using Nei's coefficient of population differentiation. Similarly, only 5% of the total diversity was attributed to differences among groups by the analysis of molecular variance (AMOVA). This small but significant difference was explained by the fact that the Central Asia group had a lower genetic variation than the other diversity centres studied.

Conclusion

We found that our sesame collection was genetically very variable and did not show an association between geographical origin and AFLP patterns. This result suggests that there was considerable gene flow among diversity centres. Future germplasm collection strategies should focus on sampling a large number of plants. Covering many diversity centres is less important because each centre represents a major part of the total diversity in sesame, Central Asia centre being the only exception. The same recommendation holds for the choice of parents for segregant populations used in breeding projects. The traditional assumption that selecting genotypes of different geographical origin will maximize the diversity available to a breeding project does not hold in sesame.     

Rapid knot detection and application to protein structure prediction

MOTIVATION: Knots in polypeptide chains have been found in very few proteins, and consequently should be generally avoided in protein structure prediction methods. Most effective structure prediction methods do not model the protein folding process itself, but rather seek only to correctly obtain the final native state. Consequently, the mechanisms that prevent knots from occurring in native proteins are not relevant to the modeling process, and as a result, knots can occur with significantly higher frequency in protein models. Here we describe Knotfind, a simple algorithm for knot detection that is fast enough for structure prediction, where tens or hundreds of thousands of conformations may be sampled during the course of a prediction. We have used this algorithm to characterize knots in large populations of model structures generated for targets in CASP 5 and CASP 6 using the Rosetta homology-based modeling method. RESULTS: Analysis of CASP5 models suggested several possible avenues for introduction of knots into these models, and these insights were applied to structure prediction in CASP 6, resulting in a significant decrease in the proportion of knotted models generated. Additionally, using the knot detection algorithm on structures in the Protein Data Bank, a previously unreported deep trefoil knot was found in acetylornithine transcarbamylase. AVAILABILITY: The Knotfind algorithm is available in the Rosetta structure prediction program at http://www.rosettacommons.org.     

A novel Arabidopsis-Colletotrichum pathosystem for the molecular dissection of plant-fungal interactions

The ability of a Colletotrichum sp., originally isolated from Brassica campestris, to infect Arabidopsis thaliana was examined. Sequence analysis of the internal transcribed spacer (ITS)1, 5.8S RNA gene and ITS2 regions of ribosomal (r)DNA showed the pathogen to be Colletotrichum destructivum. The host range was broad, including many cruciferous plants and some legumes. At 25 degrees C, all A. thaliana accessions tested were susceptible to the Brassica isolates of C. destructivum; however, at 15 degrees C, the accession Ws-2 showed a temperature-dependant resistance, in which single epidermal cells underwent a rapid hypersensitive response. Legume isolates of C. destructivum were unable to infect A. thaliana and induced deposition of callose papillae at sites of attempted penetration. In compatible interactions, C. destructivum showed a two-stage, hemibiotrophic infection process. The initial biotrophic phase was associated with large, intracellular primary hyphae and was confined to one epidermal cell; whereas, in the subsequent necrotrophic phase, narrow secondary hyphae extensively colonized the tissue and conidia were produced in acervuli. An efficient transformation system was established for C. destructivum, using Agrobacterium-mediated transfer of DNA. The ability to genetically manipulate both partners in the interaction is an important advantage, and the Arabidopsis-Colletotrichum pathosystem should provide a valuable new model for dissecting plant-fungal interactions.     

Capsaicin Treatment Attenuates Cholangiocarcinoma Carcinogenesis

Capsaicin, the most abundant pungent molecule produced by pepper plants, represents an important ingredient in spicy foods consumed throughout the world. Studies have shown that capsaicin can relieve inflammation and has anti-proliferative effects on various human malignancies. Cholangiocarcinoma (CC) is a cancer disease with rising incidence. The prognosis remains dismal with little advance in treatment. The aim of the present study is to explore the anti-tumor activity of capsaicin in cultured human CC cell lines. Capsaicin effectively impaired cell proliferation, migration, invasion, epithelial to mesenchymal transition and growth of softagar colonies. Further, we show that capsaicin treatment of CC cells regulates the Hedgehog signaling pathway. Conclusion: Our results provide a basis for capsaicin to improve the prognosis of CCs in vivo and present new insights into the effectiveness and mode of action of capsaicin.