Large-scale analysis of orthologs and paralogs under covarion-like and constant-but-different models of amino acid evolution.

Functional divergence between homologous proteins is expected to affect amino acid sequences in two main ways, which can be considered as proxies of biochemical divergence: a "covarion-like" pattern of correlated changes in evolutionary rates, and switches in conserved residues ("conserved but different"). Although these patterns have been used in case studies, a large-scale analysis is needed to estimate their frequency and distribution. We use a phylogenomic framework of animal genes to answer three questions: 1) What is the prevalence of such patterns? 2) Can we link such patterns at the amino acid level with selection inferred at the codon level? 3) Are patterns different between paralogs and orthologs? We find that covarion-like patterns are more frequently detected than "constant but different," but that only the latter are correlated with signal for positive selection. Finally, there is no obvious difference in patterns between orthologs and paralogs.

A novel ABCG-like transporter of Trypanosoma cruziis involved in natural resistance to benznidazole

Benznidazole (BZ) is one of the two drugs used for Chagas disease treatment. Nevertheless therapeutic failures of BZ have been reported, which were mostly attributed to variable drug susceptibility among Trypanosoma cruzistrains. ATP-binding cassette (ABC) transporters are involved in a variety of translocation processes and some members have been implicated in drug resistance. Here we report the characterisation of the first T. cruzi ABCG transporter gene, named TcABCG1, which is over-expressed in parasite strains naturally resistant to BZ. Comparison ofTcABCG1 gene sequence of two TcI BZ-resistant strains with CL Brener BZ-susceptible strain showed several single nucleotide polymorphisms, which determined 11 amino acid changes. CL Brener transfected with TcI transporter genes showed 40-47% increased resistance to BZ, whereas no statistical significant increment in drug resistance was observed when CL Brener was transfected with the homologous gene. Only in the parasites transfected with TcI genes there was 2-2.6-fold increased abundance of TcABCG1transporter protein. The analysis in wild type strains also suggests that the level of TcABCG1transporter is related to BZ natural resistance. The characteristics of untranslated regions of TcABCG1genes of BZ-susceptible and resistant strains were investigated by computational tools.

Modelling of Mercury’s surface composition and remote detection from the orbit with the BepiColombo Mercury Planetary Orbiter

It can be assumed that the composition of Mercury’s thin gas envelope (exosphere) is related to thecomposition of the planets crustal materials. If this relationship is true, then inferences regarding the bulkchemistry of the planet might be made from a thorough exospheric study. The most vexing of allunsolved problems is the uncertainty in the source of each component. Historically, it has been believedthat H and He come primarily from the solar wind, while Na and K originate from volatilized materialspartitioned between Mercury’s crust and meteoritic impactors. The processes that eject atoms andmolecules into the exosphere of Mercury are generally considered to be thermal vaporization, photonstimulateddesorption (PSD), impact vaporization, and ion sputtering. Each of these processes has its owntemporal and spatial dependence. The exosphere is strongly influenced by Mercury’s highly ellipticalorbit and rapid orbital speed. As a consequence the surface undergoes large fluctuations in temperatureand experiences differences of insolation with longitude. We will discuss these processes but focus moreon the expected surface composition and solar wind particle sputtering which releases material like Caand other elements from the surface minerals and discuss the relevance of composition modelling

Differential effects of selective HDAC inhibitors on macrophage inflammatory responses to the Toll-like receptor 4 agonist LPS.

Broad-spectrum inhibitors of HDACs are therapeutic in many inflammatory disease models but exacerbated disease in a mouse model of atherosclerosis. HDAC inhibitors have anti- and proinflammatory effects on macrophages in vitro. We report here that several broad-spectrum HDAC inhibitors, including TSA and SAHA, suppressed the LPS-induced mRNA expression of the proinflammatory mediators Edn-1, Ccl-7/MCP-3, and Il-12p40 but amplified the expression of the proatherogenic factors Cox-2 and Pai-1/serpine1 in primary mouse BMM. Similar effects were also apparent in LPS-stimulated TEPM and HMDM. The pro- and anti-inflammatory effects of TSA were separable over a concentration range, implying that individual HDACs have differential effects on macrophage inflammatory responses. The HDAC1-selective inhibitor, MS-275, retained proinflammatory effects (amplification of LPS-induced expression of Cox-2 and Pai-1 in BMM) but suppressed only some inflammatory responses. In contrast, 17a (a reportedly HDAC6-selective inhibitor) retained anti-inflammatory but not proinflammatory properties. Despite this, HDAC6(-/-) macrophages showed normal LPS-induced expression of HDAC-dependent inflammatory genes, arguing that the anti-inflammatory effects of 17a are not a result of inhibition of HDAC6 alone. Thus, 17a provides a tool to identify individual HDACs with proinflammatory properties.

DNA prime-protein boost based vaccination with a conserved region of leptospiral immunoglobulin-like A and B proteins enhances protection against leptospirosis

Leptospirosis is a zoonotic disease caused by pathogenic spirochetes of theLeptospira genus. Vaccination with bacterins has severe limitations. Here, we evaluated the N-terminal region of the leptospiral immunoglobulin-like B protein (LigBrep) as a vaccine candidate against leptospirosis using immunisation strategies based on DNA prime-protein boost, DNA vaccine, and subunit vaccine. Upon challenge with a virulent strain ofLeptospira interrogans, the prime-boost and DNA vaccine approaches induced significant protection in hamsters, as well as a specific IgG antibody response and sterilising immunity. Although vaccination with recombinant fragment of LigBrep also produced a strong antibody response, it was not immunoprotective. These results highlight the potential of LigBrep as a candidate antigen for an effective vaccine against leptospirosis and emphasise the use of the DNA prime-protein boost as an important strategy for vaccine development.

Contribution of efflux pumps, porins, and β-lactamases to multidrug resistance in clinical isolates of Acinetobacter baumannii.

We investigated the mechanisms of resistance to carbapenems, aminoglycosides, glycylcyclines, tetracyclines, and quinolones in 90 multiresistant clinical strains of Acinetobacter baumannii isolated from two genetically unrelated A. baumannii clones: clone PFGE-ROC-1 (53 strains producing the OXA-58 β-lactamase enzyme and 18 strains with the OXA-24 β-lactamase) and clone PFGE-HUI-1 (19 strains susceptible to carbapenems). We used real-time reverse transcriptase PCR to correlate antimicrobial resistance (MICs) with expression of genes encoding chromosomal β-lactamases (AmpC and OXA-51), porins (OmpA, CarO, Omp33, Dcap-like, OprB, Omp25, OprC, OprD, and OmpW), and proteins integral to six efflux systems (AdeABC, AdeIJK, AdeFGH, CraA, AbeM, and AmvA). Overexpression of the AdeABC system (level of expression relative to that by A. baumannii ATCC 17978, 30- to 45-fold) was significantly associated with resistance to tigecycline, minocycline, and gentamicin and other biological functions. However, hyperexpression of the AdeIJK efflux pump (level of expression relative to that by A. baumannii ATCC 17978, 8- to 10-fold) was significantly associated only with resistance to tigecycline and minocycline (to which the TetB efflux system also contributed). TetB and TetA(39) efflux pumps were detected in clinical strains and were associated with resistance to tetracyclines and doxycycline. The absence of the AdeABC system and the lack of expression of other mechanisms suggest that tigecycline-resistant strains of the PFGE-HUI-1 clone may be associated with a novel resistance-nodulation-cell efflux pump (decreased MICs in the presence of the inhibitor Phe-Arg β-naphthylamide dihydrochloride) and the TetA(39) system.

A functional and structural study of the major metalloprotease secreted by the pathogenic fungus Aspergillus fumigatus.

Fungalysins are secreted fungal peptidases with the ability to degrade the extracellular matrix proteins elastin and collagen and are thought to act as virulence factors in diseases caused by fungi. Fungalysins constitute a unique family among zinc-dependent peptidases that bears low sequence similarity to known bacterial peptidases of the thermolysin family. The crystal structure of the archetype of the fungalysin family, Aspergillus fumigatus metalloprotease (AfuMep), has been obtained for the first time. The 1.8 Å resolution structure of AfuMep corresponds to that of an autoproteolyzed proenzyme with separate polypeptide chains corresponding to the N-terminal prodomain in a binary complex with the C-terminal zinc-bound catalytic domain. The prodomain consists of a tandem of cystatin-like folds whose C-terminal end is buried into the active-site cleft of the catalytic domain. The catalytic domain harbouring the key catalytic zinc ion and its ligands, two histidines and one glutamic acid, undergoes a conspicuous rearrangement of its N-terminal end during maturation. One key positively charged amino-acid residue and the C-terminal disulfide bridge appear to contribute to its structural-functional properties. Thus, structural, biophysical and biochemical analysis were combined to provide a deeper comprehension of the underlying properties of A. fumigatus fungalysin, serving as a framework for the as yet poorly known metallopeptidases from pathogenic fungi.

Predicting response and survival in chemotherapy-treated triple-negative breast cancer.

BACKGROUND In this study, we evaluated the ability of gene expression profiles to predict chemotherapy response and survival in triple-negative breast cancer (TNBC). METHODS Gene expression and clinical-pathological data were evaluated in five independent cohorts, including three randomised clinical trials for a total of 1055 patients with TNBC, basal-like disease (BLBC) or both. Previously defined intrinsic molecular subtype and a proliferation signature were determined and tested. Each signature was tested using multivariable logistic regression models (for pCR (pathological complete response)) and Cox models (for survival). Within TNBC, interactions between each signature and the basal-like subtype (vs other subtypes) for predicting either pCR or survival were investigated. RESULTS Within TNBC, all intrinsic subtypes were identified but BLBC predominated (55-81%). Significant associations between genomic signatures and response and survival after chemotherapy were only identified within BLBC and not within TNBC as a whole. In particular, high expression of a previously identified proliferation signature, or low expression of the luminal A signature, was found independently associated with pCR and improved survival following chemotherapy across different cohorts. Significant interaction tests were only obtained between each signature and the BLBC subtype for prediction of chemotherapy response or survival. CONCLUSIONS The proliferation signature predicts response and improved survival after chemotherapy, but only within BLBC. This highlights the clinical implications of TNBC heterogeneity, and suggests that future clinical trials focused on this phenotypic subtype should consider stratifying patients as having BLBC or not.

Detection of possible variant forms of hemoglobin in HbA1c measurements by chromatography liquid ion exchange high resolution (HPLC)

Background: The glycosylated hemoglobin (HbA1c) is used to help monitor the degree of a diabetic’s hyperglycemia. Security and accuracy of the methods used in its detection are affected by variants forms of Hb or elevations in levels of Fetal Hb (HbF). These interference are the result of a change in the haemoglobin total net charge of the variant due of a substitution of one amino acid in the remaining amino terminal of the beta chain. International Standardization for HbA1c values (NGSP) not include interference assessment as part of the certification program. Therefore, the effect of each variant or the lifting of the HbF on HbA1c result should be examined in each sample depending on the detected variant and the method used for the detection of the same. The objectives were: to describe the possible variants of Hb and their interference in HbA1c measurement by our method, after the implementation of a computer program for their detection. To identify some variants detected by chromatography liquid ion exchange high resolution (HPLC) with DNA molecular sequencing.

Synthesis of bone-like structured foams

Here we present a processing route to produce multi-structured ceramic foams based on the combination of particle-stabilized foams with polymeric sponges to produce positive and negative templating structures. Polyester sponges are infiltrated with freshly produced calcium aluminate alumina foams and upon sintering either positive templating structures are produced when wetting the sponges, or negative templating foams with a percolating pore network are obtained when completely filling the sponges. Additionally, by combining different layers of these particle-stabilized foam infiltrated sponges, various different structures can be produced, including sandwich structures, pore size gradients, and ceramic bone-like structures applying to different types of bone. The particle-stabilized foams used were in situ self-hardening calcium aluminate cement enriched alumina foams to obtain crack-free samples with pore interconnections and tailorable pore sizes.

Delta-like 4 is the essential, nonredundant ligand for Notch1 during thymic T cell lineage commitment.

Thymic T cell lineage commitment is dependent on Notch1 (N1) receptor-mediated signaling. Although the physiological ligands that interact with N1 expressed on thymic precursors are currently unknown, in vitro culture systems point to Delta-like 1 (DL1) and DL4 as prime candidates. Using DL1- and DL4-lacZ reporter knock-in mice and novel monoclonal antibodies to DL1 and DL4, we show that DL4 is expressed on thymic epithelial cells (TECs), whereas DL1 is not detected. The function of DL4 was further explored in vivo by generating mice in which DL4 could be specifically inactivated in TECs or in hematopoietic progenitors. Although loss of DL4 in hematopoietic progenitors did not perturb thymus development, inactivation of DL4 in TECs led to a complete block in T cell development coupled with the ectopic appearance of immature B cells in the thymus. These immature B cells were phenotypically indistinguishable from those developing in the thymus of conditional N1 mutant mice. Collectively, our results demonstrate that DL4 is the essential and nonredundant N1 ligand responsible for T cell lineage commitment. Moreover, they strongly suggest that N1-expressing thymic progenitors interact with DL4-expressing TECs to suppress B lineage potential and to induce the first steps of intrathymic T cell development.

Circulating insulin-like growth factor-I in pregnancy and maternal risk of breast cancer.

BACKGROUND: Elevated serum concentrations of insulin-like growth factor (IGF)-1 have been associated with increased risk of breast cancer. Previously, we reported a similar association in samples obtained during pregnancy. The current study was conducted to further characterize the association of IGF-1 during pregnancy with maternal breast cancer risk. METHODS: A case-control study was nested within the Finnish Maternity Cohort. The study was limited to primiparous women less than 40 years of age, who donated blood samples during early (median, 12 weeks) pregnancy and delivered a single child at term. Seven hundred and nineteen women with invasive breast cancer were eligible. Two controls (n = 1,434) were matched to each case on age and date at blood donation. Serum IGF-1 concentration was measured using an Immulite 2000 analyzer. Conditional logistic regression was used to estimate odds ratios (OR) and 95% confidence intervals (CI).RESULTS: No significant associations were observed between serum IGF-1 concentrations and breast cancer risk in both the overall analysis (OR 1.08 (95% CI 0.80-1.47) and in analyses stratified by histological subtype, lag-time to cancer diagnosis, age at pregnancy or age at diagnosis.CONCLUSIONS: There was no association between IGF-1 and maternal breast cancer risk during early pregnancy in this large nested case-control study.Impact:Serum IGF-1 concentrations during early pregnancy may not be related to maternal risk of breast cancer.

Implications of the serpentine phase transition on the behaviour of beryllium and lithium-boron of subducted ultramafic rocks

The Totalp-Platta-Malenco ophiolites in the Eastern Central Alps offer a unique opportunity to study the behaviour of Li, Be and B in ultramafic rocks in response to serpentinization and to progressive Alpine metamorphism. These units represent the remnants of a former ocean-continent transition that was intensely serpentinized during exposure on the Jurassic seafloor of the Ligurian Tethys. From north to the south, three isograd reactions (lizardite double right arrow antigorite + brucite; lizardite + talc double right arrow antigorite; lizardite + tremolite double right arrow antigorite + diopside) have been used to quantify the evolution of the light element content of metamorphic minerals. We determined the Li, Be and B concentrations in major silicate minerals from the ultramafic bodies of Totalp, Platta and Malenco by secondary ion mass spectrometry. Mantle minerals have Be concentrations (e.g. <0.001-0.009 mu g/g in olivine) similar to the metamorphic minerals that replace them (e.g. <0.001-0.016 mu g/g in serpentine). The mantle signature of Be is thus neither erased during seafloor alteration nor by progressive metamorphism from prehnite-pumpellyite to epidote-amphibolite facies. In contrast, the Li and B inventories of metamorphic minerals are related to the lizardite-to-antigorite transition. Both elements display higher concentrations in the low-temperature serpentine polymorph lizardite (max. 156 mu/g Li, max. 318 mu g/g B) than in antigorite (max. 0.11 mu g/g Li, max. 12 mu g/g B). Calculated average B/Li ratios for lizardite (similar to 1395) and antigorite (similar to 115) indicate that Li fractionates from B during the lizardite-to-antigorite transition during prograde metamorphism in ultramafic rocks. In subduction zones, this signature is likely to be recorded in the B-rich nature of forearc fluids. Relative to oceanic mantle the Be content of mantle clinopyroxene is much higher, but similar to Be values from mantle xenoliths and subduction-related peridotite massifs. These data support previous hypothesis that the mantle rocks from the Eastern Central Alps have a subcontinental origin. We conclude that Be behaves conservatively during subduction metamorphism of ultramafic rocks, at least at low-temperature, and thus retains the fingerprint of ancient subduction-related igneous events in mantle peridotites. (C) 2010 Elsevier Ltd. All rights reserved.