CLOCK expression identifies developing circadian oscillator neurons in the brains of Drosophila embryos

<p>Abstract</p> <p>Background</p> <p>The <it>Drosophila </it>circadian oscillator is composed of transcriptional feedback loops in which CLOCK-CYCLE (CLK-CYC) heterodimers activate their feedback regulators <it>period </it>(<it>per</it>) and <it>timeless </it>(<it>tim</it>) via E-box mediated transcription. These feedback loop oscillators are present in distinct clusters of dorsal and lateral neurons in the adult brain, but how this pattern of expression is established during development is not known. Since CLK is required to initiate feedback loop function, defining the pattern of CLK expression in embryos and larvae will shed light on oscillator neuron development.</p> <p>Results</p> <p>A novel CLK antiserum is used to show that CLK expression in the larval CNS and adult brain is limited to circadian oscillator cells. CLK is initially expressed in presumptive small ventral lateral neurons (s-LN_vs), dorsal neurons 2 s (DN₂s), and dorsal neuron 1 s (DN₁s) at embryonic stage (ES) 16, and this CLK expression pattern persists through larval development. PER then accumulates in all CLK-expressing cells except presumptive DN₂s during late ES 16 and ES 17, consistent with the delayed accumulation of PER in adult oscillator neurons and antiphase cycling of PER in larval DN₂s. PER is also expressed in non-CLK-expressing cells in the embryonic CNS starting at ES 12. Although PER expression in CLK-negative cells continues in <it>Clk</it>^Jrkembryos, PER expression in cells that co-express PER and CLK is eliminated.</p> <p>Conclusion</p> <p>These data demonstrate that brain oscillator neurons begin development during embryogenesis, that PER expression in non-oscillator cells is CLK-independent, and that oscillator phase is an intrinsic characteristic of brain oscillator neurons. These results define the temporal and spatial coordinates of factors that initiate <it>Clk </it>expression, imply that circadian photoreceptors are not activated until the end of embryogenesis, and suggest that PER functions in a different capacity before oscillator cell development is initiated.</p

The Drosophila FMRP and LARK RNA-binding proteins function together to regulate eye development and circadian behavior

Fragile X syndrome (FXS) is the most common form of hereditary mental retardation. FXS patients have a deficit for the fragile X mental retardation protein (FMRP) that results in abnormal neuronal dendritic spine morphology and behavioral phenotypes, including sleep abnormalities. In a Drosophila model of FXS, flies lacking the dfmr1 protein (dFMRP) have abnormal circadian rhythms apparently as a result of altered clock output. In this study, we present biochemical and genetic evidence that dFMRP interacts with a known clock output component, the LARK RNA-binding protein. Our studies demonstrate physical interactions between dFMRP and LARK, that the two proteins are present in a complex in vivo, and that LARK promotes the stability of dFMRP. Furthermore, we show genetic interactions between the corresponding genes indicating that dFMRP and LARK function together to regulate eye development and circadian behavior

A prospective cohort study of the long-term effects of CPAP on carotid artery intima-media thickness in Obstructive sleep apnea syndrome

<p>Abstract</p> <p>Objective</p> <p>To examine the long-term effect of CPAP on carotid artery intima-media thickness (IMT) in patients with Obstructive sleep apnea syndrome(OSAS).</p> <p>Methods</p> <p>A prospective observational study over 12 months at a teaching hospital on 50 patients newly diagnosed with OSAS who received CPAP or conservative treatment (CT). Carotid IMT was assessed with B-mode Doppler ultrasound from both carotid arteries using images of the far wall of the distal 10 mm of the common carotid arteries at baseline, 6 months and 12 months.</p> <p>Measurements and results [mean (SE)]</p> <p>Altogether 28 and 22 patients received CPAP and CT respectively without significant differences in age 48.8(1.8) vs 50.5(2.0)yrs, BMI 28.2(0.7) vs 28.0(1.2)kg/m2, ESS 13.1(0.7) vs 12.7(0.6), AHI 38(3) vs 39(3)/hr, arousal index 29(2) vs 29(2)/hr, minimum SaO₂75(2) vs 77(2)% and existing co-morbidities. CPAP usage was 4.6(0.3) and 4.7(0.4)hrs/night over 6 months and 1 year respectively. Carotid artery IMT at baseline, 6 months, and 12 months were 758(30), 721(20), and 705(20)micron for the CPAP group versus 760(30), 770(30), and 778(30)micron respectively for the CT group, p = 0.002.</p> <p>Among those free of cardiovascular disease(n = 24), the carotid artery IMT at baseline, 6 months and 12 months were 722(40), 691(40), and 659(30)micron for the CPAP group (n = 12) with usage 4.5(0.7) and 4.7(0.7) hrs/night over 6 months and 12 months whereas the IMT data for the CT group(n = 12) were 660(20), 685(10), and 690(20)micron respectively, p = 0.006.</p> <p>Conclusions</p> <p>Reduction of carotid artery IMT occurred mostly in the first 6 months and was sustained at 12 months in patients with reasonable CPAP compliance.</p

Phosphorylation of a Central Clock Transcription Factor Is Required for Thermal but Not Photic Entrainment

Transcriptional/translational feedback loops drive daily cycles of expression in clock genes and clock-controlled genes, which ultimately underlie many of the overt circadian rhythms manifested by organisms. Moreover, phosphorylation of clock proteins plays crucial roles in the temporal regulation of clock protein activity, stability and subcellular localization. dCLOCK (dCLK), the master transcription factor driving cyclical gene expression and the rate-limiting component in the Drosophila circadian clock, undergoes daily changes in phosphorylation. However, the physiological role of dCLK phosphorylation is not clear. Using a Drosophila tissue culture system, we identified multiple phosphorylation sites on dCLK. Expression of a mutated version of dCLK where all the mapped phospho-sites were switched to alanine (dCLK-15A) rescues the arrythmicity of Clk(out) flies, yet with an approximately 1.5 hr shorter period. The dCLK-15A protein attains substantially higher levels in flies compared to the control situation, and also appears to have enhanced transcriptional activity, consistent with the observed higher peak values and amplitudes in the mRNA rhythms of several core clock genes. Surprisingly, the clock-controlled daily activity rhythm in dCLK-15A expressing flies does not synchronize properly to daily temperature cycles, although there is no defect in aligning to light/dark cycles. Our findings suggest a novel role for clock protein phosphorylation in governing the relative strengths of entraining modalities by adjusting the dynamics of circadian gene expression

Roadmap on commercialization of metal halide perovskite photovoltaics

Perovskite solar cells (PSCs) represent one of the most promising emerging photovoltaic technologies due to their high power conversion efficiency. However, despite the huge progress made not only in terms of the efficiency achieved, but also fundamental understanding of the relevant physics of the devices and issues which affect their efficiency and stability, there are still unresolved problems and obstacles on the path toward commercialization of this promising technology. In this roadmap, we aim to provide a concise and up to date summary of outstanding issues and challenges, and the progress made toward addressing these issues. While the format of this article is not meant to be a comprehensive review of the topic, it provides a collection of the viewpoints of the experts in the field, which covers a broad range of topics related to PSC commercialization, including those relevant for manufacturing (scaling up, different types of devices), operation and stability (various factors), and environmental issues (in particular the use of lead). We hope that the article will provide a useful resource for researchers in the field and that it will facilitate discussions and move forward toward addressing the outstanding challenges in this fast-developing field

Functioning of the dimeric GABA(B) receptor extracellular domain revealed by glycan wedge scanning

The G-protein-coupled receptor (GPCR) activated by the neurotransmitter GABA is made up of two subunits, GABA(B1) and GABA(B2). GABA(B1) binds agonists, whereas GABA(B2) is required for trafficking GABA(B1) to the cell surface, increasing agonist affinity to GABA(B1), and activating associated G proteins. These subunits each comprise two domains, a Venus flytrap domain (VFT) and a heptahelical transmembrane domain (7TM). How agonist binding to the GABA(B1) VFT leads to GABA(B2) 7TM activation remains unknown. Here, we used a glycan wedge scanning approach to investigate how the GABA(B) VFT dimer controls receptor activity. We first identified the dimerization interface using a bioinformatics approach and then showed that introducing an N-glycan at this interface prevents the association of the two subunits and abolishes all activities of GABA(B2), including agonist activation of the G protein. We also identified a second region in the VFT where insertion of an N-glycan does not prevent dimerization, but blocks agonist activation of the receptor. These data provide new insight into the function of this prototypical GPCR and demonstrate that a change in the dimerization interface is required for receptor activation

Tuning the energetics and tailoring the optical properties of silver clusters confined in zeolites

The integration of metal atoms and clusters in well-defined dielectric cavities is a powerful strategy to impart new properties to them that depend on the size and geometry of the confined space as well as on metal-host electrostatic interactions. Here, we unravel the dependence of the electronic properties of metal clusters on space confinement by studying the ionization potential of silver clusters embedded in four different zeolite environments over a range of silver concentrations. Extensive characterization reveals a strong influence of silver loading and host environment on the cluster ionization potential, which is also correlated to the cluster's optical and structural properties. Through fine-tuning of the zeolite host environment, we demonstrate photoluminescence quantum yields approaching unity. This work extends our understanding of structure property relationships of small metal clusters and applies this understanding to develop highly photoluminescent materials with potential applications in optoelectronics and bioimaging