Control of TRPM3 Ion Channels by Protein Kinase CK2-Mediated Phosphorylation in Pancreatic β-Cells of the Line INS-1

In pancreatic β-cells of the line INS-1, glucose uptake and metabolism induce the openings of Ca2+-permeable TRPM3 channels that contribute to the elevation of the intracellular Ca2+ concen tration and the fusion of insulin granules with the plasma membrane. Conversely, glucose-induced Ca2+ signals and insulin release are reduced by the activity of the serine/threonine kinase CK2. There fore, we hypothesized that TRPM3 channels might be regulated by CK2 phosphorylation. We used recombinant TRPM3α2 proteins, native TRPM3 proteins from INS-1 β-cells, and TRPM3-derived oligopeptides to analyze and localize CK2-dependent phosphorylation of TRPM3 channels. The func tional consequences of CK2 phosphorylation upon TRPM3-mediated Ca2+ entry were investigated in Fura-2 Ca2+-imaging experiments. Recombinant TRPM3α2 channels expressed in HEK293 cells displayed enhanced Ca2+ entry in the presence of the CK2 inhibitor CX-4945 and their activity was strongly reduced after CK2 overexpression. TRPM3α2 channels were phosphorylated by CK2 in vitro at serine residue 1172. Accordingly, a TRPM3α2 S1172A mutant displayed enhanced Ca2+ entry. The TRPM3-mediated Ca2+ entry in INS-1 β-cells was also strongly increased in the presence of CX-4945 and reduced after overexpression of CK2. Our study shows that CK2-mediated phosphorylation controls TRPM3 channel activity in INS-1 β-cells

Atomic structures of fibrillar segments of hIAPP suggest tightly mated β-sheets are important for cytotoxicity.

hIAPP fibrils are associated with Type-II Diabetes, but the link of hIAPP structure to islet cell death remains elusive. Here we observe that hIAPP fibrils are cytotoxic to cultured pancreatic β-cells, leading us to determine the structure and cytotoxicity of protein segments composing the amyloid spine of hIAPP. Using the cryoEM method MicroED, we discover that one segment, 19-29 S20G, forms pairs of β-sheets mated by a dry interface that share structural features with and are similarly cytotoxic to full-length hIAPP fibrils. In contrast, a second segment, 15-25 WT, forms non-toxic labile β-sheets. These segments possess different structures and cytotoxic effects, however, both can seed full-length hIAPP, and cause hIAPP to take on the cytotoxic and structural features of that segment. These results suggest that protein segment structures represent polymorphs of their parent protein and that segment 19-29 S20G may serve as a model for the toxic spine of hIAPP

Understanding and Controlling Cu-Catalyzed Graphene Nucleation: The Role of Impurities, Roughness, and Oxygen Scavenging

The mechanism by which Cu catalyst pretreatments control graphene nucleation density in scalable chemical vapor deposition (CVD) is systematically explored. The intrinsic and extrinsic carbon contamination in the Cu foil is identified by time-of-flight secondary ion mass spectrometry as a major factor influencing graphene nucleation and growth. By selectively oxidizing the backside of the Cu foil prior to graphene growth, a drastic reduction of the graphene nucleation density by 6 orders of magnitude can be obtained. This approach decouples surface roughness effects and at the same time allows us to trace the scavenging effect of oxygen on deleterious carbon impurities as it permeates through the Cu bulk. Parallels to well-known processes in Cu metallurgy are discussed. We also put into context the relative effectiveness and underlying mechanisms of the most widely used Cu pretreatments, including wet etching and electropolishing, allowing a rationalization of current literature and determination of the relevant parameter space for graphene growth. Taking into account the wider CVD growth parameter space, guidelines are discussed for high-throughput manufacturing of "electronic-quality" monolayer graphene films with domain size exceeding 1 mm, suitable for emerging industrial applications, such as electronics and photonics.This research was supported by the ERC under grant InsituNANO (279342), the EPSRC under grant GRAPHTED (EP/K016636/1), and the Innovation R&D programme of the National Measurement System of the U.K. Department of Business, Innovation and Skills (project number 118616)

CK2 Activity Mediates the Aggressive Molecular Signature of Glioblastoma Multiforme by Inducing Nerve/Glial Antigen (NG)2 Expression

Nerve/glial antigen (NG)2 expression crucially determines the aggressiveness of glioblastoma multiforme (GBM). Recent evidence suggests that protein kinase CK2 regulates NG2 expression. Therefore, we investigated in the present study whether CK2 inhibition suppresses proliferation and migration of NG2-positive GBM cells. For this purpose, CK2 activity was suppressed in the NG2-positive cell lines A1207 and U87 by the pharmacological inhibitor CX-4945 and CRISPR/Cas9- mediated knockout of CK2α. As shown by quantitative real-time PCR, luciferase-reporter assays, flow cytometry and western blot, this significantly reduced NG2 gene and protein expression when compared to vehicle-treated and wild type controls. In addition, CK2 inhibition markedly reduced NG2-dependent A1207 and U87 cell proliferation and migration. The Cancer Genome Atlas (TCGA)- based data further revealed not only a high expression of both NG2 and CK2 in GBM but also a positive correlation between the mRNA expression of the two proteins. Finally, we verified a decreased NG2 expression after CX-4945 treatment in patient-derived GBM cells. These findings indicate that the inhibition of CK2 represents a promising approach to suppress the aggressive molecular signature of NG2-positive GBM cells. Therefore, CX-4945 may be a suitable drug for the future treatment of NG2-positive GBM

Transient Receptor Potential Vanilloid 6 (TRPV6) Proteins Control the Extracellular Matrix Structure of the Placental Labyrinth

Calcium-selective transient receptor potential Vanilloid 6 (TRPV6) channels are expressed in fetal labyrinth trophoblasts as part of the feto–maternal barrier, necessary for sufficient calcium supply, embryo growth, and bone development during pregnancy. Recently, we have shown a less- compact labyrinth morphology of Trpv6-deficient placentae, and reduced Ca2+ uptake of primary trophoblasts upon functional deletion of TRPV6. Trpv6-/- trophoblasts show a distinct calcium-dependent phenotype. Deep proteomic profiling of wt and Trpv6-/- primary trophoblasts using label-free quantitative mass spectrometry leads to the identification of 2778 proteins. Among those, a group of proteases, including high-temperature requirement A serine peptidase 1 (HTRA1) and different granzymes are more abundantly expressed in Trpv6-/- trophoblast lysates, whereas the extracellular matrix protein fibronectin and the fibronectin-domain-containing protein 3A (FND3A) were markedly reduced. Trpv6-/- placenta lysates contain a higher intrinsic proteolytic activity increasing fibronectin degradation. Our results show that the extracellular matrix formation of the placental labyrinth depends on TRPV6; its deletion in trophoblasts correlates with the increased expression of proteases controlling the extracellular matrix in the labyrinth during pregnancy

Contactless graphene conductivity mapping on a wide range of substrates with terahertz time-domain reflection spectroscopy.

We demonstrate how terahertz time-domain spectroscopy (THz-TDS) operating in reflection geometry can be used for quantitative conductivity mapping of large area chemical vapour deposited graphene films on sapphire, silicon dioxide/silicon and germanium. We validate the technique against measurements performed with previously established conventional transmission based THz-TDS and are able to resolve conductivity changes in response to induced back-gate voltages. Compared to the transmission geometry, measurement in reflection mode requires careful alignment and complex analysis, but circumvents the need of a terahertz transparent substrate, potentially enabling fast, contactless, in-line characterisation of graphene films on non-insulating substrates such as germanium.H.L. and J.A.Z. acknowledge financial support from the EPSRC (Grant No. EP/L019922/1). P.B.W. acknowledges EPSRC Cambridge NanoDTC EP/G037221/1. R.D., H.E.B. and D. R. acknowledge financial support from the EPSRC (Grant No. EP/J017671/1, Coherent Terahertz Systems). S.H. acknowledges funding from the EPSRC (Grant No. EP/K016636/1, GRAPHTED)

Hot subdwarf binaries - Masses and nature of their heavy compact companions

Neutron stars and stellar-mass black holes are the remnants of massive stars, which ended their lives in supernova explosions. These exotic objects can only be studied in relatively rare cases. If they are interacting with close companions they become bright X-ray sources. If they are neutron stars, they may be detected as pulsars. Only a few hundred such systems are presently known in the Galaxy. However, there should be many more binaries with basically invisible compact objects in non-interacting binaries. Here we report the discovery of unseen compact companions to hot subdwarfs in close binary systems. Hot subdwarfs are evolved helium-core-burning stars that have lost most of their hydrogen envelopes, often due to binary interactions. Using high-resolution spectra and assuming tidal synchronisation of the subdwarfs, we were able to constrain the companion masses of 32 binaries. While most hot subdwarf binaries have white-dwarf or late-type main sequence companions, as predicted by binary evolution models, at least 5% of the observed subdwarfs must have very massive companions: unusually heavy white dwarfs, neutron stars and, in some cases, even black holes. We present evolutionary models which show that such binaries can indeed form if the system has evolved through two common-envelope phases. This new connection between hot subdwarfs, which are numerous in the Galaxy, and massive compact objects may lead to a tremendous increase in the number of known neutron stars and black holes and shed some light on this dark population and its evolutionary link to the X-ray binary population.Comment: 8 pages, 5 figures, to appear in the Journal of Physics Conference Proceedings (JPCS) for the 16th European White Dwarf Workshop, Barcelona, Spain, June 30 - July 11, 200

Prolactin-sensitive olfactory sensory neurons regulate male preference in female mice by modulating responses to chemosensory cues

Chemosensory cues detected in the nose need to be integrated with the hormonal status to trigger appropriate behaviors, but the neural circuits linking the olfactory and the endocrine system are insufficiently understood. Here, we characterize olfactory sensory neurons in the murine nose that respond to the pituitary hormone prolactin. Deletion of prolactin receptor in these cells results in impaired detection of social odors and blunts male preference in females. The prolactin-responsive olfactory sensory neurons exhibit a distinctive projection pattern to the brain that is similar across different individuals and express a limited subset of chemosensory receptors. Prolactin modulates the responses within these neurons to discrete chemosensory cues contained in male urine, providing a mechanism by which the hormonal status can be directly linked with distinct olfactory cues to generate appropriate behavioral responses

TRPM3 Is a Nociceptor Channel Involved in the Detection of Noxious Heat

SummaryTransient receptor potential melastatin-3 (TRPM3) is a broadly expressed Ca2+-permeable nonselective cation channel. Previous work has demonstrated robust activation of TRPM3 by the neuroactive steroid pregnenolone sulfate (PS), but its in vivo gating mechanisms and functions remained poorly understood. Here, we provide evidence that TRPM3 functions as a chemo- and thermosensor in the somatosensory system. TRPM3 is molecularly and functionally expressed in a large subset of small-diameter sensory neurons from dorsal root and trigeminal ganglia, and mediates the aversive and nocifensive behavioral responses to PS. Moreover, we demonstrate that TRPM3 is steeply activated by heating and underlies heat sensitivity in a subset of sensory neurons. TRPM3-deficient mice exhibited clear deficits in their avoidance responses to noxious heat and in the development of inflammatory heat hyperalgesia. These experiments reveal an unanticipated role for TRPM3 as a thermosensitive nociceptor channel implicated in the detection of noxious heat

A systems approach towards remote health-monitoring in older adults: Introducing a zero-interaction digital exhaust.

Using connected sensing devices to remotely monitor health is a promising way to help transition healthcare from a rather reactive to a more precision medicine oriented proactive approach, which could be particularly relevant in the face of rapid population ageing and the challenges it poses to healthcare systems. Sensor derived digital measures of health, such as digital biomarkers or digital clinical outcome assessments, may be used to monitor health status or the risk of adverse events like falls. Current research around such digital measures has largely focused on exploring the use of few individual measures obtained through mobile devices. However, especially for long-term applications in older adults, this choice of technology may not be ideal and could further add to the digital divide. Moreover, large-scale systems biology approaches, like genomics, have already proven beneficial in precision medicine, making it plausible that the same could also hold for remote-health monitoring. In this context, we introduce and describe a zero-interaction digital exhaust: a set of 1268 digital measures that cover large parts of a person's activity, behavior and physiology. Making this approach more inclusive of older adults, we base this set entirely on contactless, zero-interaction sensing technologies. Applying the resulting digital exhaust to real-world data, we then demonstrate the possibility to create multiple ageing relevant digital clinical outcome assessments. Paired with modern machine learning, we find these assessments to be surprisingly powerful and often on-par with mobile approaches. Lastly, we highlight the possibility to discover novel digital biomarkers based on this large-scale approach