Visualizing and Quantifying Intracellular Behavior and Abundance of the Core Circadian Clock Protein PERIOD2

SummaryTranscriptional-translational feedback loops (TTFLs) are a conserved molecular motif of circadian clocks. The principal clock in mammals is the suprachiasmatic nucleus (SCN) of the hypothalamus. In SCN neurons, auto-regulatory feedback on core clock genes Period (Per) and Cryptochrome (Cry) following nuclear entry of their protein products is the basis of circadian oscillation [1, 2]. In Drosophila clock neurons, the movement of dPer into the nucleus is subject to a circadian gate that generates a delay in the TTFL, and this delay is thought to be critical for oscillation [3, 4]. Analysis of the Drosophila clock has strongly influenced models of the mammalian clock, and such models typically infer complex spatiotemporal, intracellular behaviors of mammalian clock proteins. There are, however, no direct measures of the intracellular behavior of endogenous circadian proteins to support this: dynamic analyses have been limited and often have no circadian dimension [5–7]. We therefore generated a knockin mouse expressing a fluorescent fusion of native PER2 protein (PER2::VENUS) for live imaging. PER2::VENUS recapitulates the circadian functions of wild-type PER2 and, importantly, the behavior of PER2::VENUS runs counter to the Drosophila model: it does not exhibit circadian gating of nuclear entry. Using fluorescent imaging of PER2::VENUS, we acquired the first measures of mobility, molecular concentration, and localization of an endogenous circadian protein in individual mammalian cells, and we showed how the mobility and nuclear translocation of PER2 are regulated by casein kinase. These results provide new qualitative and quantitative insights into the cellular mechanism of the mammalian circadian clock

The clock genes Period 2 and Cryptochrome 2 differentially balance bone formation

Background: Clock genes and their protein products regulate circadian rhythms in mammals but have also been implicated in various physiological processes, including bone formation. Osteoblasts build new mineralized bone whereas osteoclasts degrade it thereby balancing bone formation. To evaluate the contribution of clock components in this process, we investigated mice mutant in clock genes for a bone volume phenotype. Methodology/Principal Findings: We found that Per2Brdm1 mutant mice as well as mice lacking Cry2-/- displayed significantly increased bone volume at 12 weeks of age, when bone turnover is high. Per2Brdm1 mutant mice showed alterations in parameters specific for osteoblasts whereas mice lacking Cry2-/- displayed changes in osteoclast specific parameters. Interestingly, inactivation of both Per2 and Cry2 genes leads to normal bone volume as observed in wild type animals. Importantly, osteoclast parameters affected due to the lack of Cry2, remained at the level seen in the Cry2-/- mutants despite the simultaneous inactivation of Per2. Conclusions/Significance: This indicates that Cry2 and Per2 affect distinct pathways in the regulation of bone volume with Cry2 influencing mostly the osteoclastic cellular component of bone and Per2 acting on osteoblast parameters

Brief Report: Association of Tumor Necrosis Factor Receptor–Associated Periodic Syndrome With Gonosomal Mosaicism of a Novel 24-Nucleotide TNFRSF1A Deletion

Objective: To investigate the molecular cause of persistent fevers in a patient returning from working overseas, in whom investigations for tropical diseases yielded negative results. / Methods: DNA was extracted from the patient's whole blood, leukocyte subpopulations, saliva, hair root, and sperm. The TNFRSF1A gene was analyzed by polymerase chain reaction (PCR), allele-specific PCR, Sanger sequencing, and next-generation sequencing. In silico molecular modeling was performed to predict the structural and functional consequences of the tumor necrosis factor receptor (TNFR) type I protein mutation in the extracellular domain. / Results: Sanger sequencing corroborated by allele-specific PCR detected a novel in-frame deletion of 24 nucleotides (c.255_278del) in the TNFRSF1A gene, and this was subsequently confirmed using next-generation sequencing methods (targeted sequencing and amplicon-based deep sequencing). Results of amplicon-based deep sequencing revealed variable frequency of the mutant allele among different cell lines, including sperm, thus supporting the presence of gonosomal TNFRSF1A mosaicism. The patient had a complete response to treatment with interleukin-1 (IL-1) blockade, with resolution of symptoms and normalization of acute-phase protein levels. / Conclusion: We describe the first case of gonosomal TNFRSF1A mosaicism in a patient with TNFR-associated periodic syndrome (TRAPS), which was attributable to a novel, somatic 24-nucleotide in-frame deletion. The clinical picture in this patient, including the complete response to IL-1 blockade, was typical of that found in TRAPS. This case adds TRAPS to the list of dominantly inherited autoinflammatory diseases reported to be caused by somatic (or postzygotic) mutation

The HEV Ventilator

HEV is a low-cost, versatile, high-quality ventilator, which has been designed in response to the COVID-19 pandemic. The ventilator is intended to be used both in and out of hospital intensive care units, and for both invasive and non-invasive ventilation. The hardware can be complemented with an external turbine for use in regions where compressed air supplies are not reliably available. The standard modes provided include PC-A/C(Pressure Assist Control),PC-A/C-PRVC(Pressure Regulated Volume Control), PC-PSV (Pressure Support Ventilation) and CPAP (Continuous Positive airway pressure). HEV is designed to support remote training and post market surveillance via a web interface and data logging to complement the standard touch screen operation, making it suitable for a wide range of geographical deployment. The HEV design places emphasis on the quality of the pressure curves and the reactivity of the trigger, delivering a global performance which will be applicable to ventilator needs beyond theCOVID-19 pandemic. This article describes the conceptual design and presents the prototype units together with their performance evaluation.Comment: 34 pages, 18 figures, Extended version of the article submitted to PNA

Close-range remote sensing of Saturn's rings during Cassini's ring-grazing orbits and Grand Finale

Saturn’s rings are an accessible exemplar of an astrophysical disk, tracing the Saturn system’s dynamical processes and history. We present close-range remote-sensing observations of the main rings from the Cassini spacecraft. We find detailed sculpting of the rings by embedded masses, and banded texture belts throughout the rings. Saturn-orbiting streams of material impact the F ring. There are fine-scaled correlations among optical depth, spectral properties, and temperature in the B ring, but anticorrelations within strong density waves in the A ring. There is no spectral distinction between plateaux and the rest of the C ring, whereas the region outward of the Keeler gap is spectrally distinct from nearby regions. These results likely indicate that radial stratification of particle physical properties, rather than compositional differences, is responsible for producing these ring structures. © 2019 American Association for the Advancement of Science. All rights reserved

Circadian Rhythm and Sleep Disruption: Causes, Metabolic Consequences and Countermeasures.

Circadian (∼ 24 hour) timing systems pervade all kingdoms of life, and temporally optimize behaviour and physiology in humans. Relatively recent changes to our environments, such as the introduction of artificial lighting, can disorganize the circadian system, from the level of the molecular clocks that regulate the timing of cellular activities to the level of synchronization between our daily cycles of behaviour and the solar day. Sleep/wake cycles are intertwined with the circadian system, and global trends indicate that these too are increasingly subject to disruption. A large proportion of the world's population is at increased risk of environmentally-driven circadian rhythm and sleep disruption, and a minority of individuals are also genetically predisposed to circadian misalignment and sleep disorders. The consequences of disruption to the circadian system and sleep are profound and include myriad metabolic ramifications, some of which may be compounded by adverse effects on dietary choices. If not addressed, the deleterious effects of such disruption will continue to cause widespread health problems; therefore, implementation of the numerous behavioural and pharmaceutical interventions that can help restore circadian system alignment and enhance sleep will be important

Cassini CIRS and ISS opposition effects of Saturn's rings -I. C ring narrow or broad surge?

International audienceWe focus on the thermal and optical opposition effects of Saturn's C ring seen by Cassini CIRS (Composite InfraRed Spectrometer) at 15.7 μm and ISS (Imaging Science Subsystem) at 0.6 μm. The opposition surge is a brightness peak observed at low phase angle (α → 0 •). Saturn rings' opposition surge was recently observed in reflected light and thermal infrared emission by Cassini. There is debate on whether the C ring's thermal opposition surge width is narrow (1 •) or broad (30 •). This surge is important because its width was used to define the scale of ring properties driving the thermal peak. We parametrize the CIRS and ISS phase curves with several morphological models to fit the surge shape. For five of the largest C ring's plateaus, we find that their thermal surge is 10 times wider than the optical surge and that the thermal surge width (∼4 •) is neither narrow, nor broad. We compare radial differences between CIRS and ISS surge morphologies with the optical depth τ (from UVIS, UltraViolet Imaging Spectrograph) and water ice band depth (from VIMS, Visual and Infrared Mapping Spectrometer) profiles. We find that: water ice band depths (microscopic ring signatures) and τ (macroscopic ring signatures) show respectively little and large contrasts between the background and the plateaus. The thermal surge amplitude and τ are correlated, and we found no band depth dependence, contrary to the optical surge amplitude, which shows no correlation with τ. These correlations suggest a macroscopic scale dominance in controlling the C ring's thermal opposition effect

Temperature Variations of Saturn Rings with Viewing Geometries from Prime to Equinox Cassini Missions

After more than six years in orbit around Saturn, the Cassini Composite Infrared Spectrometer (CIRS) has acquired an extensive set of measurements of Saturn's main rings (A, B, C and Cassini Division) in the thermal infrared. Temperatures were retrieved for the lit and unlit rings over a variety of ring geometries that include phase angle, solar and spacecraft elevations and local time. We show that some of these parameters (solar and spacecraft elevations, phase angle) play a role in the temperature variations in the first order, while the others (ring and particle local time) produced second order effects. The results of this comparison will be presented

Two numerical models designed to reproduce Saturn ring temperatures as measured by Cassini-CIRS

International audienc