Restoration of mutant cytochrome P450 reductase activity by external flavin

Cytochrome P450 oxidoreductase (POR) supplies electrons from NADPH to steroid and drug metabolizing reactions catalyzed by the cytochrome P450s located in endoplasmic reticulum. Mutations in human POR cause a wide spectrum of disease ranging from disordered steroidogenesis to sexual differentiation. Previously we and others have shown that POR mutations can lead to reduced activities of steroidogenic P450s CYP17A1, CYP19A1 and CYP21A1. Here we are reporting that mutations in the FMN binding domain of POR may reduce CYP3A4 activity, potentially influencing drug and steroid metabolism; and the loss of CYP3A4 activity may be correlated to the reduction of cytochrome b(5) by POR. Computational molecular docking experiments with a FMN free structural model of POR revealed that an external FMN could be docked in close proximity to the FAD moiety and receive electrons donated by NADPH. Using FMN supplemented assays we have demonstrated restoration of the defective POR activity in vitro.

NADPH-cytochrome P450 oxidoreductase: roles in physiology, pharmacology, and toxicology

This is a report on a symposium sponsored by the American Society for Pharmacology and Experimental Therapeutics and held at the Experimental Biology 2012 meeting in San Diego, California, on April 25, 2012. The symposium speakers summarized and critically evaluated our current understanding of the physiologic, pharmacological, and toxicological roles of NADPH-cytochrome P450 oxidoreductase (POR), a flavoprotein involved in electron transfer to microsomal cytochromes P450 (P450), cytochrome b(5), squalene mono-oxygenase, and heme oxygenase. Considerable insight has been derived from the development and characterization of mouse models with conditional Por deletion in particular tissues or partial suppression of POR expression in all tissues. Additional mouse models with global or conditional hepatic deletion of cytochrome b(5) are helping to clarify the P450 isoform- and substrate-specific influences of cytochrome b(5) on P450 electron transfer and catalytic function. This symposium also considered studies using siRNA to suppress POR expression in a hepatoma cell-culture model to explore the basis of the hepatic lipidosis phenotype observed in mice with conditional deletion of Por in liver. The symposium concluded with a strong translational perspective, relating the basic science of human POR structure and function to the impacts of POR genetic variation on human drug and steroid metabolism.

Hybridization between distant lineages increases adaptive variation during a biological invasion: stickleback in Switzerland

The three-spined stickleback is a widespread Holarctic species complex that radiated from the sea into freshwaters after the retreat of the Pleistocene ice sheets. In Switzerland, sticklebacks were absent with the exception of the far northwest, but different introduced populations have expanded to occupy a wide range of habitats since the late 19th century. A well-studied adaptive phenotypic trait in sticklebacks is the number of lateral plates. With few exceptions, freshwater and marine populations in Europe are fixed for either the low plated phenotype or the fully plated phenotype, respectively. Switzerland, in contrast, harbours in close proximity the full range of phenotypic variation known from across the continent. We addressed the phylogeographic origins of Swiss sticklebacks using mitochondrial partial cytochrome b and control region sequences. We found only five different haplotypes but these originated from three distinct European regions, fixed for different plate phenotypes. These lineages occur largely in isolation at opposite ends of Switzerland, but co-occur in a large central part. Across the country, we found a strong correlation between a microsatellite linked to the high plate ectodysplasin allele and the mitochondrial haplotype from a region where the fully plated phenotype is fixed. Phylogenomic and population genomic analysis of 481 polymorphic amplified fragment length polymorphism loci indicate genetic admixture in the central part of the country. The same part of the country also carries elevated within-population phenotypic variation. We conclude that during the recent invasive range expansion of sticklebacks in Switzerland, adaptive and neutral between-population genetic variation was converted into within-population variation, raising the possibility that hybridization between colonizing lineages contributed to the ecological success of sticklebacks in Switzerland.

Divergent evolutionary processes associated with colonization of offshore islands

Oceanic islands have been a test ground for evolutionary theory, but here, we focus on the possibilities for evolutionary study created by offshore islands. These can be colonized through various means and by a wide range of species, including those with low dispersal capabilities. We use morphology, modern and ancient sequences of cytochrome b (cytb) and microsatellite genotypes to examine colonization history and evolutionary change associated with occupation of the Orkney archipelago by the common vole (Microtus arvalis), a species found in continental Europe but not in Britain. Among possible colonization scenarios, our results are most consistent with human introduction at least 5100 bp (confirmed by radiocarbon dating). We used approximate Bayesian computation of population history to infer the coast of Belgium as the possible source and estimated the evolutionary timescale using a Bayesian coalescent approach. We showed substantial morphological divergence of the island populations, including a size increase presumably driven by selection and reduced microsatellite variation likely reflecting founder events and genetic drift. More surprisingly, our results suggest that a recent and widespread cytb replacement event in the continental source area purged cytb variation there, whereas the ancestral diversity is largely retained in the colonized islands as a genetic ‘ark’. The replacement event in the continental M. arvalis was probably triggered by anthropogenic causes (land-use change). Our studies illustrate that small offshore islands can act as field laboratories for studying various evolutionary processes over relatively short timescales, informing about the mainland source area as well as the island.

Differential diagnosis of the honey bee trypanosomatids Crithidia mellificae and Lotmaria passim

Trypanosomatids infecting honey bees have been poorly studied with molecular methods until recently. After the description of Crithidia mellificae (Langridge and McGhee, 1967) it took about forty years until molecular data for honey bee trypanosomatids became available and were used to identify and describe a new trypanosomatid species from honey bees, Lotmaria passim (Evans and Schwarz, 2014). However, an easy method to distinguish them without sequencing is not yet available. Research on the related bumble bee parasites Crithidia bombi and Crithidia expoeki revealed a fragment length polymorphism in the internal transcribed spacer 1 (ITS1), which enabled species discrimination. In search of fragment length polymorphisms for differential diagnostics in honey bee trypanosomatids, we studied honey bee trypanosomatid cell cultures of C. mellificae and L. passim. This research resulted in the identification of fragment length polymorphisms in ITS1 and ITS1-2 markers, which enabled us to develop a diagnostic method to differentiate both honey bee trypanosomatid species without the need for sequencing. However, the amplification success of the ITS1 marker depends probably on the trypanosomatid infection level. Further investigation confirmed that L. passim is the dominant species in Belgium, Japan and Switzerland. We found C. mellificae only rarely in Belgian honey bee samples, but not in honey bee samples from other countries. C. mellificae was also detected in mason bees (Osmia bicornis and Osmia cornuta) besides in honey bees. Further, the characterization and comparison of additional markers from L. passim strain SF (published as C. mellificae strain SF) and a Belgian honey bee sample revealed very low divergence in the 18S rRNA, ITS1-2, 28S rRNA and cytochrome b sequences. Nevertheless, a variable stretch was observed in the gp63 virulence factor.

Search for pair production of heavy top-like quarks decaying to a high-pT W boson and a b quark in the lepton plus jets final state at sqrts=7 TeV with the ATLAS detector

A search is presented for production of a heavy up-type quark (t') together with its antiparticle, assuming a significant branching ratio for subsequent decay into a W boson and a b quark. The search is based on 4.7 fb(-1) of pp collisions root s = 7 TeV recorded in 2011 with the ATLAS detector at the CERN Large Hadron Collider. Data are analyzed in the lepton + jets final state, characterized by a high-transverse-momentum isolated electron or muon, large missing transverse momentum and at least three jets. The analysis strategy relies on the substantial boost of the W bosons in the t'(t') over bar signal when m(t') greater than or similar to 400 GeV. No significant excess of events above the Standard Model expectation is observed and the result of the search is interpreted in the context of fourth-generation and vector-like quark models. Under the assumption of a branching ratio BR(t' -> W b) = I, a fourth-generation t' quark with mass lower than 656 GeV is excluded at 95% confidence level. In addition, in light of the recent discovery of a new boson of mass similar to 126 GeV at the LHC, upper limits are derived in the two-dimensional plane of BR(t' -> Wb) versus BR(t' -> Ht), where H is the Standard Model Higgs boson, for vector-like quarks of various masses.

Search for a heavy particle decaying into an electron and a muon with the ATLAS detector in sqrt[s] = 7 TeV pp collisions at the LHC

This Letter presents the first search for a heavy particle decaying into an e ± μ(-/+) final state in sqrt[s] = 7 TeV pp collisions at the LHC. The data were recorded by the ATLAS detector during 2010 and correspond to a total integrated luminosity of 35 pb(-1). No excess above the standard model background expectation is observed. Exclusions at 95% confidence level are placed on two representative models. In an R-parity violating supersymmetric model, tau sneutrinos with a mass below 0.75 TeV are excluded, assuming all R-parity violating couplings are zero except λ(311)' = 0.11 and λ312 = 0.07. In a lepton flavor violating model, a Z'-like vector boson with masses of 0.70-1.00 TeV and corresponding cross sections times branching ratios of 0.175-0.183 pb is excluded. These results extend to higher mass R-parity violating sneutrinos and lepton flavor violating Z's than previous constraints from the Tevatron.

Search for a heavy narrow resonance decaying to eμ, eτ, or μτ with the ATLAS detector in View the MathML sourcepp collisions at the LHC

This Letter presents the results of a search for a heavy particle decaying into an e(+/-)mu(+/-), e(+/-)tau(+/-), or mu(+/-)tau(+/-) final state in pp collisions at root s = 7 TeV. The data were recorded with the ATLAS detector at the LHC during 2011 and correspond to an integrated luminosity of 4.6 fb(-1). No significant excess above the Standard Model expectation is observed, and exclusions at 95% confidence level are placed on the cross section times branching ratio for the production of an R-parity-violating supersymmetric tau sneutrino. For a sneutrino mass of 500 (2000) GeV, the observed limits on the production cross section times branching ratio are 3.2 (1.4) fb, 42 (17) fb, and 40 (18) fb for the e mu, e tau, and mu tau modes, respectively. These results considerably extend constraints from Tevatron experiments.