Phototropins do not alter accumulation of evening-phased circadian transcripts under blue light.

The circadian system induces rhythmic variation in a suite of biochemical and physiological processes that serves to optimise plant growth in diel cycles. To be of greatest utility, these rhythmic behaviours are coordinated with regular environmental changes such as the rising and setting of the sun. Photoreceptors, and metabolites produced during photosynthesis, act to synchronise the internal timing mechanism with lighting cues. We have recently shown that phototropins help maintain robust rhythms of photosynthetic operating efficiency (?PSII or Fq'/Fm') under blue light, although rhythmic accumulation of morning-phased circadian transcripts in the nucleus was unaffected. Here we report that evening-phased nuclear clock transcripts were also unaffected. We also observe that rhythms of nuclear clock transcript accumulation are maintained in phototropin mutant plants under a fluctuating lighting regime that induced a loss of Fq'/Fm' rhythms

Caspase 8 activation independent of Fas (CD95/APO-1) signaling may mediate killing of B-chronic lymphocytic leukemia cells by cytotoxic drugs or gamma radiation.

Ligation of the cell-surface Fas molecule by its ligand (Fas-L) or agonistic anti-Fas monoclonal antibodies results in the cleavage and activation of the cysteine protease procaspase 8 followed by the activation of procaspase 3 and by apoptosis. In some leukemia cell lines, cytotoxic drugs induce expression of Fas-L, which may contribute to cell killing through the ligation of Fas. The involvement of Fas, Fas-L, and caspase 8 was studied in the killing of B-cell chronic lymphocytic leukemia (B-CLL) cells by chlorambucil, fludarabine, or gamma radiation. Spontaneous apoptosis was observed at 24-hour incubation, with additional apoptosis induced by each of the cytotoxic treatments. Although Fas mRNA expression was elevated after exposure to chlorambucil, fludarabine, or gamma radiation, Fas protein levels only increased after irradiation. Therefore, Fas expression may be regulated by multiple mechanisms that allow the translation of Fas mRNA only in response to restricted cytotoxic stimuli. None of the cytotoxic stimuli studied here induced Fas-L expression. An agonistic anti-Fas monoclonal antibody (CH-11) did not significantly augment apoptosis induction by any of the death stimuli. A Fas-blocking antibody (ZB4) did not inhibit spontaneous, chlorambucil-, fludarabine-, or radiation-induced apoptosis. However, procaspase 8 processing was induced by all cytotoxic stimuli. These data suggest that the Fas/Fas-L signaling system does not play a major role in the induction of apoptosis in B-CLL cells treated with cytotoxic drugs or radiation. However, Fas-independent activation of caspase 8 may play a crucial role in the regulation of apoptosis in these cells

Disrupted hippocampal sharp-wave ripple-associated spike dynamics in a transgenic mouse model of dementia.

This is the peer reviewed version of the article, which has been published in final form at DOI: 10.1113/jphysiol.2014.282889. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.Neurons within the CA1 region of the hippocampus are co-activated during high frequency (100-250 Hz) sharp wave ripple (SWR) activity in a manner that likely drives synaptic plasticity and promotes memory consolidation. In this study we have used a transgenic mouse model of dementia (rTg4510 mice) which overexpresses a mutant form of tau protein, to examine the effects of tauopathy on hippocampal SWRs and associated neuronal firing. Tetrodes were used to record simultaneous extracellular action potentials and local field potentials from the dorsal CA1 pyramidal cell layer of 7-8 month old wild-type and rTg4510 mice at rest in their home cage. At this age point these mice exhibit neurofibrillary tangles, neurodegeneration and cognitive deficits. Epochs of sleep or quiet restfulness were characterised by minimal locomotor activity and a low theta/delta ratio in the local field potential power spectrum. SWRs detected off-line were significantly lower in amplitude and had an altered temporal structure in rTg4510 mice. Nevertheless, the average frequency profile and duration of the SWRs were relatively unaltered. Putative interneurons displayed significantly less temporal and phase locking to SWRs in rTg4510 mice, whilst putative pyramidal neurons showed increased temporal and phase locking to SWRs. These findings indicate there is reduced inhibitory control of hippocampal network events and points to a novel mechanism which may contribute to impairments in memory consolidation in this model of dementia. This article is protected by copyright. All rights reserved.Alzheimer’s Research UKMedical Research Counci

Geldanamycin and herbimycin A induce apoptotic killing of B chronic lymphocytic leukemia cells and augment the cells' sensitivity to cytotoxic drugs.

We studied the actions of geldanamycin (GA) and herbimycin A (HMA), inhibitors of the chaperone proteins Hsp90 and GRP94, on B chronic lymphocytic leukemia (CLL) cells in vitro. Both drugs induced apoptosis of the majority of CLL isolates studied. Whereas exposure to 4-hour pulses of 30 to 100 nM GA killed normal B lymphocytes and CLL cells with similar dose responses, T lymphocytes from healthy donors as well as those present in the CLL isolates were relatively resistant. GA, but not HMA, showed a modest cytoprotective effect toward CD34+ hematopoietic progenitors from normal bone marrow. The ability of bone marrow progenitors to form hematopoietic colonies was unaffected by pulse exposures to GA. Both GA and HMA synergized with chlorambucil and fludarabine in killing a subset of CLL isolates. GA- and HMA-induced apoptosis was preceded by the up-regulation of the stress-responsive chaperones Hsp70 and BiP. Both ansamycins also resulted in down-regulation of Akt protein kinase, a modulator of cell survival. The relative resistance of T lymphocytes and of CD34+ bone marrow progenitors to GA coupled with its ability to induce apoptosis following brief exposures and to synergize with cytotoxic drugs warrant further investigation of ansamycins as potential therapeutic agents in CLL

Solution structure of a repeated unit of the ABA-1 nematode polyprotein allergen of ascaris reveals a novel fold and two discrete lipid-binding sites

Parasitic nematode worms cause serious health problems in humans and other animals. They can induce allergic-type immune responses, which can be harmful but may at the same time protect against the infections. Allergens are proteins that trigger allergic reactions and these parasites produce a type that is confined to nematodes, the nematode polyprotein allergens (NPAs). These are synthesized as large precursor proteins comprising repeating units of similar amino acid sequence that are subsequently cleaved into multiple copies of the allergen protein. NPAs bind small lipids such as fatty acids and retinol (Vitamin A) and probably transport these sensitive and insoluble compounds between the tissues of the worms. Nematodes cannot synthesize these lipids, so NPAs may also be crucial for extracting nutrients from their hosts. They may also be involved in altering immune responses by controlling the lipids by which the immune and inflammatory cells communicate. We describe the molecular structure of one unit of an NPA, the well-known ABA-1 allergen of Ascaris and find its structure to be of a type not previously found for lipid-binding proteins, and we describe the unusual sites where lipids bind within this structur

Altered intrinsic pyramidal neuron properties and pathway-specific synaptic dysfunction underlie aberrant hippocampal network function in a mouse model of Tauopathy.

Final published version of article. This article is freely available online through the J Neurosci Author Open Choice option.The formation and deposition of tau protein aggregates is proposed to contribute to cognitive impairments in dementia by disrupting neuronal function in brain regions, including the hippocampus. We used a battery of in vivo and in vitro electrophysiological recordings in the rTg4510 transgenic mouse model, which overexpresses a mutant form of human tau protein, to investigate the effects of tau pathology on hippocampal neuronal function in area CA1 of 7- to 8-month-old mice, an age point at which rTg4510 animals exhibit advanced tau pathology and progressive neurodegeneration. In vitro recordings revealed shifted theta-frequency resonance properties of CA1 pyramidal neurons, deficits in synaptic transmission at Schaffer collateral synapses, and blunted plasticity and imbalanced inhibition at temporoammonic synapses. These changes were associated with aberrant CA1 network oscillations, pyramidal neuron bursting, and spatial information coding in vivo. Our findings relate tauopathy-associated changes in cellular neurophysiology to altered behavior-dependent network function. SIGNIFICANCE STATEMENT: Dementia is characterized by the loss of learning and memory ability. The deposition of tau protein aggregates in the brain is a pathological hallmark of dementia; and the hippocampus, a brain structure known to be critical in processing learning and memory, is one of the first and most heavily affected regions. Our results show that, in area CA1 of hippocampus, a region involved in spatial learning and memory, tau pathology is associated with specific disturbances in synaptic, cellular, and network-level function, culminating in the aberrant encoding of spatial information and spatial memory impairment. These studies identify several novel ways in which hippocampal information processing may be disrupted in dementia, which may provide targets for future therapeutic intervention.Medical Research Council (MRC)Royal SocietyAlzheimer's Research United Kingdo

Examining the Relationships among the Coaching Climate, Life Skills Development and Well-Being in Sport

Using Benson and Saito's framework for positive youth development, we investigated the relationships between the coaching climate, young people's perceived life skills development within sport, and their psychological well-being. British youth sport participants (N=326, Mage=13.81, range=11–18 years) completed a survey assessing the coaching climate, participants' perceived life skills development (teamwork, goal setting, time management, emotional skills, interpersonal communication, social skills, leadership, and problem solving and decision making) and psychological well-being (self-esteem, positive affect, and satisfaction with life). In all analyses, the coaching climate was positively related to young peoples' perceived development of life skills within sport and their psychological well-being. Total life skills development (a summative score of all eight life skills scores) was positively related to all three psychological well-being indicators – providing support for the "pile-up" effect – and partially mediated the relationships between the coaching climate and participants' psychological well-being. Interpretation of the results indicated that coaches should foster the development of multiple life skills in youth sport participants, as they are associated with participants' psychological well-being. One way this can be achieved is through autonomy-supportive coaching behaviours

Determinants of the voltage dependence of G protein modulation within calcium channel β subunits

CaVβ subunits of voltage-gated calcium channels contain two conserved domains, a src-homology-3 (SH3) domain and a guanylate kinase-like (GK) domain with an intervening HOOK domain. We have shown in a previous study that, although Gβγ-mediated inhibitory modulation of CaV2.2 channels did not require the interaction of a CaVβ subunit with the CaVα1 subunit, when such interaction was prevented by a mutation in the α1 subunit, G protein modulation could not be removed by a large depolarization and showed voltage-independent properties (Leroy et al., J Neurosci 25:6984–6996, 2005). In this study, we have investigated the ability of mutant and truncated CaVβ subunits to support voltage-dependent G protein modulation in order to determine the minimal domain of the CaVβ subunit that is required for this process. We have coexpressed the CaVβ subunit constructs with CaV2.2 and α2δ-2, studied modulation by the activation of the dopamine D2 receptor, and also examined basal tonic modulation. Our main finding is that the CaVβ subunit GK domains, from either β1b or β2, are sufficient to restore voltage dependence to G protein modulation. We also found that the removal of the variable HOOK region from β2a promotes tonic voltage-dependent G protein modulation. We propose that the absence of the HOOK region enhances Gβγ binding affinity, leading to greater tonic modulation by basal levels of Gβγ. This tonic modulation requires the presence of an SH3 domain, as tonic modulation is not supported by any of the CaVβ subunit GK domains alone

Evaluation of alternative mosquito sampling methods for malaria vectors in Lowland South - East Zambia.

Sampling malaria vectors and measuring their biting density is of paramount importance for entomological surveys of malaria transmission. Human landing catch (HLC) has been traditionally regarded as a gold standard method for surveying human exposure to mosquito bites. However, due to the risk of human participant exposure to mosquito-borne parasites and viruses, a variety of alternative, exposure-free trapping methods were compared in lowland, south-east Zambia. Centres for Disease Control and Prevention miniature light trap (CDC-LT), Ifakara Tent Trap model C (ITT-C), resting boxes (RB) and window exit traps (WET) were all compared with HLC using a 3 × 3 Latin Squares design replicated in 4 blocks of 3 houses with long lasting insecticidal nets, half of which were also sprayed with a residual deltamethrin formulation, which was repeated for 10 rounds of 3 nights of rotation each during both the dry and wet seasons. The mean catches of HLC indoor, HLC outdoor, CDC-LT, ITT-C, WET, RB indoor and RB outdoor, were 1.687, 1.004, 3.267, 0.088, 0.004, 0.000 and 0.008 for Anopheles quadriannulatus Theobald respectively, and 7.287, 6.784, 10.958, 5.875, 0.296, 0.158 and 0.458, for An. funestus Giles, respectively. Indoor CDC-LT was more efficient in sampling An. quadriannulatus and An. funestus than HLC indoor (Relative rate [95% Confidence Interval] = 1.873 [1.653, 2.122] and 1.532 [1.441, 1.628], respectively, P < 0.001 for both). ITT-C was the only other alternative which had comparable sensitivity (RR = 0.821 [0.765, 0.881], P < 0.001), relative to HLC indoor other than CDC-LT for sampling An. funestus. While the two most sensitive exposure-free techniques primarily capture host-seeking mosquitoes, both have substantial disadvantages for routine community-based surveillance applications: the CDC-LT requires regular recharging of batteries while the bulkiness of ITT-C makes it difficult to move between sampling locations. RB placed indoors or outdoors and WET had consistently poor sensitivity so it may be useful to evaluate additional alternative methods, such as pyrethrum spray catches and back packer aspirators, for catching resting mosquitoes

Stage-Specific Inhibition of MHC Class I Presentation by the Epstein-Barr Virus BNLF2a Protein during Virus Lytic Cycle

gamma-herpesvirus Epstein-Barr virus (EBV) persists for life in infected individuals despite the presence of a strong immune response. During the lytic cycle of EBV many viral proteins are expressed, potentially allowing virally infected cells to be recognized and eliminated by CD8+ T cells. We have recently identified an immune evasion protein encoded by EBV, BNLF2a, which is expressed in early phase lytic replication and inhibits peptide- and ATP-binding functions of the transporter associated with antigen processing. Ectopic expression of BNLF2a causes decreased surface MHC class I expression and inhibits the presentation of indicator antigens to CD8+ T cells. Here we sought to examine the influence of BNLF2a when expressed naturally during EBV lytic replication. We generated a BNLF2a-deleted recombinant EBV (ΔBNLF2a) and compared the ability of ΔBNLF2a and wild-type EBV-transformed B cell lines to be recognized by CD8+ T cell clones specific for EBV-encoded immediate early, early and late lytic antigens. Epitopes derived from immediate early and early expressed proteins were better recognized when presented by ΔBNLF2a transformed cells compared to wild-type virus transformants. However, recognition of late antigens by CD8+ T cells remained equally poor when presented by both wild-type and ΔBNLF2a cell targets. Analysis of BNLF2a and target protein expression kinetics showed that although BNLF2a is expressed during early phase replication, it is expressed at a time when there is an upregulation of immediate early proteins and initiation of early protein synthesis. Interestingly, BNLF2a protein expression was found to be lost by late lytic cycle yet ΔBNLF2a-transformed cells in late stage replication downregulated surface MHC class I to a similar extent as wild-type EBV-transformed cells. These data show that BNLF2a-mediated expression is stage-specific, affecting presentation of immediate early and early proteins, and that other evasion mechanisms operate later in the lytic cycle