The rockefeller university

Transmission of Plasmodium species from a mammalian host to the mosquito vector requires the uptake, during an infected blood meal, of gametocytes, the precursor cells of the gametes. Relatively little is known about the molecular mechanisms involved in the developmental switch from asexual development to sexual differentiation or the maturation and survival of gametocytes. Here, we show that a gene coding for a novel putative transporter, NPT1, plays a crucial role in the development of Plasmodium berghei gametocytes. Parasites lacking NPT1 are severely compromised in the production of gametocytes and the rare gametocytes produced are unable to differentiate into fertile gametes. This is the earliest block in gametocytogenesis obtained by reverse genetics and the first to demonstrate the role of a protein with a putative transport function in sexual development. These results and the high degree of conservation of NPT1 in Plasmodium species suggest that this protein could be an attractive target for the development of novel drugs to block the spread of malaria.

Background: Atopic dermatitis (AD) is a common inflammatory skin disease exhibiting a predominantly T(H)2/"T22'' immune activation and a defective epidermal barrier. Narrow-band UVB (NB-UVB) is considered an efficient treatment for moderate-to-severe AD. In patients with psoriasis, NB-UVB has been found to suppress T(H)1/T(H)17 polarization, with subsequent reversal of epidermal hyperplasia. The immunomodulatory effects of this treatment are largely unknown in patients with AD. Objective: We sought to evaluate the effects of NB-UVB on immune and barrier abnormalities in patients with AD, aiming to establish reversibility of disease and biomarkers of therapeutic response. Methods: Twelve patients with moderate-to-severe chronic AD received NB-UVB phototherapy 3 times weekly for up to 12 weeks. Lesional and nonlesional skin biopsy specimens were obtained before and after treatment and evaluated by using gene expression and immunohistochemistry studies. Results: All patients had at least a 50% reduction in SCORAD index scores with NB-UVB phototherapy. The T(H)2, T22, and T(H)1 immune pathways were suppressed, and measures of epidermal hyperplasia and differentiation normalized. The reversal of disease activity was associated with elimination of inflammatory leukocytes and T(H)2/T22-associated cytokines and chemokines and normalized expression of barrier proteins. Conclusions: Our study shows that resolution of clinical disease in patients with chronic AD is accompanied by reversal of both the epidermal defects and the underlying immune activation. We have defined a set of biomarkers of disease response that associate resolved T(H)2 and T22 inflammation in patients with chronic AD with reversal of barrier pathology. By showing reversal of the AD epidermal phenotype with a broad immune-targeted therapy, our data argue against a fixed genetic phenotype. (J Allergy Clin Immunol 2011;128:583-93.)

Cocaine induces fast dopamine increases in brain striatal regions, which are recognized to underlie its rewarding effects. Both dopamine D1 and D2 receptors are involved in cocaine's reward but the dynamic downstream consequences of cocaine effects in striatum are not fully understood. Here we used transgenic mice expressing EGFP under the control of either the D1 receptor (D1R) or the D2 receptor (D2R) gene and microprobe optical imaging to assess the dynamic changes in intracellular calcium ([Ca2+](i)) responses (used as marker of neuronal activation) to acute cocaine in vivo separately for D1R-versus D2R-expressing neurons in striatum. Acute cocaine (8 mg/kg, i. p.) rapidly increased [Ca2+](i) in D1R-expressing neurons (10.6 +/- 3.2%) in striatum within 8.3 +/- 2.3 min after cocaine administration after which the increases plateaued; these fast [Ca2+](i) increases were blocked by pretreatment with a D1R antagonist (SCH23390). In contrast, cocaine induced progressive decreases in [Ca2+](i) in D2R-expressing neurons (10.4 +/- 5.8%) continuously throughout the 30 min that followed cocaine administration; these slower [Ca2+](i) decreases were blocked by pretreatment with a D2R antagonist (raclopride). Since activation of striatal D1R-expressing neurons (direct-pathway) enhances cocaine reward, whereas activation of D2R-expressing neurons suppresses it (indirect-pathway) (Lobo et al., 2010), this suggests that cocaine's rewarding effects entail both its fast stimulation of D1R(resulting in abrupt activation of direct-pathway neurons) and a slower stimulation of D2R(resulting in longer-lasting deactivation of indirect-pathway neurons). We also provide direct in vivo evidence of D2R and D1R interactions in the striatal responses to acute cocaine administration.

We present a study of B-s(0) decays to the CP-odd final state J/psi f(0)(980) with J/psi -> mu(+)mu(-) and f(0)(980) -> pi(+)pi(-). Using p(p) over bar $ collision data with an integrated luminosity of 3.8 fb(-1) collected by the CDF II detector at the Tevatron we measure a B-s(0) lifetime of tau(B-s(0) -> J/psi f(0)(980)) = 1.70(-0.11)(+0.12)(stat) +/- 0.03(syst)ps. This is the first measurement of the B-s(0) lifetime in a decay to a CP eigenstate and corresponds in the standard model to the lifetime of the heavy B-s(0) eigenstate. We also measure the product of branching fractions of B-s(0) -> J/psi f(0)(980) and f(0)(980) -> pi(+)pi(-) relative to the product of branching fractions of B-s(0) -> J/psi phi and phi -> K+K- to be R-f0/phi = 0.257 +/- 0: 020(stat) +/- 0.014(syst), which is the most precise determination of this quantity to date.

Autosomal dominant TLR3 deficiency has been identified as a genetic etiology of childhood herpes simplex virus 1 (HSV-1) encephalitis (HSE). This defect is partial, as it results in impaired, but not abolished induction of IFN-beta and -lambda in fibroblasts in response to TLR3 stimulation. The apparently normal resistance of these patients to other infections, viral illnesses in particular, may thus result from residual TLR3 responses. We report here an autosomal recessive form of complete TLR3 deficiency in a young man who developed HSE in childhood but remained normally resistant to other infections. This patient is compound heterozygous for two loss-of-function TLR3 alleles, resulting in an absence of response to TLR3 activation by polyinosinic-polycytidylic acid (poly(I:C)) and related agonists in his fibroblasts. Moreover, upon infection of the patient's fibroblasts with HSV-1, the impairment of IFN-beta and -lambda production resulted in high levels of viral replication and cell death. In contrast, the patient's peripheral blood mononuclear cells responded normally to poly(I:C) and to all viruses tested, including HSV-1. Consistently, various TLR3-deficient leukocytes from the patient, including CD14(+) and/or CD16(+) monocytes, plasmacytoid dendritic cells, and in vitro derived monocyte-derived macrophages, responded normally to both poly(I:C) and HSV-1, with the induction of antiviral IFN production. These findings identify a new genetic etiology for childhood HSE, indicating that TLR3-mediated immunity is essential for protective immunity to HSV-1 in the central nervous system (CNS) during primary infection in childhood, in at least some patients. They also indicate that human TLR3 is largely redundant for responses to double-stranded RNA and HSV-1 in various leukocytes, probably accounting for the redundancy of TLR3 for host defense against viruses, including HSV-1, outside the CNS.

Sequencing of small RNA cDNA libraries is an important tool for the discovery of new RNAs and the analysis of their mutational status as well as expression changes across samples. It requires multiple enzyme-catalyzed steps, including sequential oligonucleotide adapter ligations to the 3' and 5' ends of the small RNAs, reverse transcription (RT), and PCR. We assessed biases in representation of miRNAs relative to their input concentration, using a pool of 770 synthetic miRNAs and 45 calibrator oligoribonucleotides, and tested the influence of Rnl1 and two variants of Rnl2, Rnl2(1-249) and Rnl2(1-249)K227Q, for 3'-adapter ligation. The use of the Rnl2 variants for adapter ligations yielded substantially fewer side products compared with Rnl1; however, the benefits of using Rnl2 remained largely obscured by additional biases in the 5'-adapter ligation step; RT and PCR steps did not have a significant impact on read frequencies. Intramolecular secondary structures of miRNA and/or miRNA/3'-adapter products contributed to these biases, which were highly reproducible under defined experimental conditions. We used the synthetic miRNA cocktail to derive correction factors for approximation of the absolute levels of individual miRNAs in biological samples. Finally, we evaluated the influence of 5'-terminal 5-nt barcode extensions for a set of 20 barcoded 3' adapters and observed similar biases in miRNA read distribution, thereby enabling cost-saving multiplex analysis for large-scale miRNA profiling.

Evolution can follow predictable genetic trajectories(1), indicating that discrete environmental shifts can select for reproducible genetic changes(2-4). Conspecific individuals are an important feature of an animal's environment, and a potential source of selective pressures. Here we show that adaptation of two Caenorhabditis species to growth at high density, a feature common to domestic environments, occurs by reproducible genetic changes to pheromone receptor genes. Chemical communication through pheromones that accumulate during high-density growth causes young nematode larvae to enter the long-lived but non-reproductive dauer stage. Two strains of Caenorhabditis elegans grown at high density have independently acquired multigenic resistance to pheromone-induced dauer formation. In each strain, resistance to the pheromone ascaroside C3 results from a deletion that disrupts the adjacent chemoreceptor genes serpentine receptor class g (srg)-36 and -37. Through misexpression experiments, we show that these genes encode redundant G-protein-coupled receptors for ascaroside C3. Multigenic resistance to dauer formation has also arisen in high-density cultures of a different nematode species, Caenorhabditis briggsae, resulting in part from deletion of an srg gene paralogous to srg-36 and srg-37. These results demonstrate rapid remodelling of the chemoreceptor repertoire as an adaptation to specific environments, and indicate that parallel changes to a common genetic substrate can affect life-history traits across species.

Proteomic studies have identified a plethora of lysine acetylated proteins in eukaryotes and bacteria. Determining the individual lysine acetyltransferases responsible for each protein acetylation mark is crucial for elucidating the underlying regulatory mechanisms, but has been challenging due to limited biochemical methods. Here, we describe the application of a bioorthogonal chemical proteomics method to profile and identify substrates of individual lysine acetyltransferases. Addition of 4-pentynoyl-coenzyme A, an alkynyl chemical reporter for protein acetylation, to cell extracts, together with purified lysine acetyltransferase p300, enabled the fluorescent profiling and identification of protein substrates via Cu(I)-catalyzed alkyne-azide cycloaddition. We identified several known protein substrates of the acetyltransferase p300 as well as the lysine residues that were modified. Interestingly, several new candidate p300 substrates and their sites of acetylation were also discovered using this approach. Our results demonstrate that bioorthogonal chemical proteomics allows the rapid substrate identification of individual protein acetyltransferases in vitro. (C) 2011 Elsevier Ltd. All rights reserved.

A search for pair-production of first generation scalar leptoquarks is performed in the final state containing an electron, a neutrino, and at least two jets using proton-proton collision data at root s = 7 TeV. The data were collected by the CMS detector at the LHC, corresponding to an integrated luminosity of 36 pb(-1). The number of observed events is in good agreement with the predictions for standard model processes. Prior CMS results in the dielectron channel are combined with this electron + neutrino search. A 95% confidence level combined lower limit is set on the mass of a first generation scalar leptoquark at 339 GeV for beta = 0.5, where beta is the branching fraction of the leptoquark to an electron and a quark. These results represent the most stringent direct limits to date for values of beta greater than 0.05. (C) 2011 CERN. Published by Elsevier B.V. All rights reserved.