Nobel Laureates of the University of California

Natural products that elicit discomfort or pain represent invaluable tools for probing molecular mechanisms underlying pain sensation. Plant-derived irritants have predominated in this regard, but animal venoms have also evolved to avert predators by targeting neurons and receptors whose activation produces noxious sensations. As such, venoms provide a rich and varied source of small molecule and protein pharmacophores that can be exploited to characterize and manipulate key components of the pain-signalling pathway. With this in mind, here we perform an unbiased in vitro screen to identify snake venoms capable of activating somatosensory neurons. Venom from the Texas coral snake (Micrurus tener tener), whose bite produces intense and unremitting pain, excites a large cohort of sensory neurons. The purified active species (MitTx) consists of a heteromeric complex between Kunitz- and phospholipase-A2-like proteins that together function as a potent, persistent and selective agonist for acid-sensing ion channels (ASICs), showing equal or greater efficacy compared with acidic pH. MitTx is highly selective for the ASIC1 subtype at neutral pH; under more acidic conditions (pH < 6.5), MitTx massively potentiates (>100-fold) proton-evoked activation of ASIC2a channels. These observations raise the possibility that ASIC channels function as coincidence detectors for extracellular protons and other, as yet unidentified, endogenous factors. Purified MitTx elicits robust pain-related behaviour in mice by activation of ASIC1 channels on capsaicin-sensitive nerve fibres. These findings reveal a mechanism whereby snake venoms produce pain, and highlight an unexpected contribution of ASIC1 channels to nociception.

When integral membrane proteins are visualized in detergents or other artificial systems, an important layer of information is lost regarding lipid interactions and their effects on protein structure. This is especially relevant to proteins for which lipid

© 2015 Macmillan Publishers Limited. All rights reserved.The TRPA1 ion channel (also known as the wasabi receptor) is a detector of noxious chemical agents encountered in our environment or produced endogenously during tissue injury or drug metabolism. The

In the version of this Article initially published, there were errors in the author affiliations, Fig. 3, main text and Acknowledgements section.

CRISPR-Cas systems provide adaptive immunity in bacteria and archaea, beginning with integration of foreign sequences into the host CRISPR genomic locus and followed by transcription and maturation of CRISPR RNAs (crRNAs). In some CRISPR systems, a reverse transcriptase (RT) fusion to the Cas1 integrase and Cas6 maturase creates a single protein that enables concerted sequence integration and crRNA production. To elucidate how the RT-integrase organizes distinct enzymatic activities, we present the cryo-EM structure of a Cas6-RT-Cas1-Cas2 CRISPR integrase complex. The structure reveals a heterohexamer in which the RT directly contacts the integrase and maturase domains, suggesting functional coordination between all three active sites. Together with biochemical experiments, our data support a model of sequential enzymatic activities that enable CRISPR sequence acquisition from RNA and DNA substrates. These findings highlight an expanded capacity of some CRISPR systems to acquire diverse sequences that direct CRISPR-mediated interference.

CRISPR-Cas systems provide bacteria and archaea with programmable immunity against mobile genetic elements. Evolutionary pressure by CRISPR-Cas has driven bacteriophage to evolve small protein inhibitors, anti-CRISPRs (Acrs), that block Cas enzyme function by wide-ranging mechanisms. We show here that the inhibitor AcrVA4 uses a previously undescribed strategy to recognize the L. bacterium Cas12a (LbCas12a) pre-crRNA processing nuclease, forming a Cas12a dimer, and allosterically inhibiting DNA binding. The Ac. species Cas12a (AsCas12a) enzyme, widely used for genome editing applications, contains an ancestral helical bundle that blocks AcrVA4 binding and allows it to escape anti-CRISPR recognition. Using biochemical, microbiological, and human cell editing experiments, we show that Cas12a orthologs can be rendered either sensitive or resistant to AcrVA4 through rational structural engineering informed by evolution. Together, these findings explain a new mode of CRISPR-Cas inhibition and illustrate how structural variability in Cas effectors can drive opportunistic co-evolution of inhibitors by bacteriophage.

Owing to its extremely high luminosity and long duration, supernova (SN) 2006gy radiated more energy in visual light than any other known SN. Two hypotheses to explain its high luminosity at early times - that it was powered by shock interaction with circumstellar material (CSM) as implied by its Type IIn spectrum, or that it was fueled by radioactive decay from a large mass of 56Ni synthesized in a pair-instability SN-predicted different late-time properties. Here we present observations of SN 2006gy obtained more than a year after discovery. We were unable to detect it at visual wavelengths, but clear near-infrared (IR) K′ and H-band detections show that it is still at least as luminous as the peak of a normal Type II SN. We also present spectra giving an upper limit to the late-time Ha luminosity of ≲S10 39 erg s-1. Based on the weak late-time Ha, X-ray, and radio emission, combined with the difficulty of explaining the shift to IR wavelengths, we can rule out ongoing CSM interaction as the primary late-time power source of SN 2006gy. Instead, we propose that the evolution of SN 2006gy is consistent with one of two possible scenarios: (1)apairinstability SN plus modest CSM interaction, where the radioactive decay luminosity shifts to the IR because of dust formation; or (2) an IR echo, where radiation emitted during peak luminosity heats a pre-existing dust shell at radii near 1 light year, requiring the progenitor star to have ejected another shell of ∼10 M ⊙ about 1500 yr before the SN. © 2008. The American Astronomical Society. All rights reserved.

Motion of short-period stars orbiting the supermassive black hole in our Galactic Center has been monitored for more than 20 years. These observations are currently offering a new way to test the gravitational theory in an unexplored regime: in a strong gravitational field, around a supermassive black hole. In this proceeding, we present three results: (i) a constraint on a hypothetical fifth force obtained by using 19 years of observations of the two best measured short-period stars S0-2 and S0-38; (ii) an upper limit on the secular advance of the argument of the periastron for the star S0-2; (iii) a sensitivity analysis showing that the relativistic redshift of S0-2 will be measured after its closest approach to the black hole in 2018.

We summarize work on the central parsec of the Galactic center based on imaging and spectroscopic observations at the Keck and Gemini telescopes. These observations include stellar positions in two dimension and the velocity in three dimensions. Spectroscopic observations also enables measurements of the physical properties of individual stars, such as the spectral type and in some cases the effective temperature, metallicity, and surface gravity. These observations show a complex stellar population with a young (4-6 Myr) compact star cluster in the central 0.5 pc embedded in in an older and much more massive nuclear star cluster. Surprisingly, the old late-type giants do not show a cusp profile as long been expected from theoretical work. The majority of the stars have higher than solar metallicity, with only about 6% of the stars having [M/Fe] < -0.5, which is consistent with an origin from the MW disk.

Large coat protein complex II (COPII)-coated vesicles serve to convey the large cargo procollagen I (PC1) from the endoplasmic reticulum (ER). The link between large cargo in the lumen of the ER and modulation of the COPII machinery remains unresolved. TANGO1 is required for PC secretion and interacts with PC and COPII on opposite sides of the ER membrane, but evidence suggests that TANGO1 is retained in the ER, and not included in normal size (<100 nm) COPII vesicles. Here we show that TANGO1 is exported out of the ER in large COPII-coated PC1 carriers, and retrieved back to the ER by the retrograde coat, COPI, mediated by the C-terminal RDEL retrieval sequence of HSP47. TANGO1 is known to target the COPII initiation factor SEC12 to ER exit sites through an interacting protein, cTAGE5. SEC12 is important for the growth of COPII vesicles, but it is not sorted into small budded vesicles. We found both cTAGE5 and SEC12 were exported with TANGO1 in large COPII carriers. In contrast to its exclusion from small transport vesicles, SEC12 was particularly enriched around ER membranes and large COPII carriers that contained PC1. We constructed a split GFP system to recapitulate the targeting of SEC12 to PC1 via the luminal domain of TANGO1. The minimal targeting system enriched SEC12 around PC1 and generated large PC1 carriers. We conclude that TANGO1, cTAGE5, and SEC12 are copacked with PC1 into COPII carriers to increase the size of COPII, thus ensuring the capture of large cargo.

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In addition to its role in forming vesicles from the endoplasmic reticulum (ER), the coat protein complex II (COPII) is also responsible for selecting specific cargo proteins to be packaged into COPII transport vesicles. Comparison of COPII vesicle formation in mammalian systems and in yeast suggested that the former uses more elaborate mechanisms for cargo recognition, presumably to cope with a significantly expanded repertoire of cargo that transits the secretory pathway. Using proTGFα, the transmembrane precursor of transforming growth factor α (TGFα), as a model cargo protein, we demonstrate in cell-free assays that at least one auxiliary cytosolic factor is specifically required for the efficient packaging of proTGFα into COPII vesicles. Using a knockout HeLa cell line generated by CRISPR/Cas9, we provide functional evidence showing that a transmembrane protein, Cornichon-1 (CNIH), acts as a cargo receptor of proTGFα. We show that both CNIH and the auxiliary cytosolic factor(s) are required for efficient recruitment of proTGFα to the COPII coat in vitro. Moreover, we provide evidence that the recruitment of cargo protein by the COPII coat precedes and may be distinct from subsequent cargo packaging into COPII vesicles.

We present the discovery and measurements of a gravitationally lensed supernova (SN) behind the galaxy cluster MOO J1014+0038. Based on multi-band Hubble Space Telescope and Very Large Telescope (VLT) photometry of the supernova, and VLT spectroscopy of the host galaxy, we find a 97.5% probability that this SN is a SN Ia, and a 2.5% chance of a CC SN. Our typing algorithm combines the shape and color of the light curve with the expected rates of each SN type in the host galaxy. With a redshift of 2.2216, this is the highest redshift SN Ia discovered with a spectroscopic host-galaxy redshift. A further distinguishing feature is that the lensing cluster, at redshift 1.23, is the most distant to date to have an amplified SN. The SN lies in the middle of the color and light-curve shape distributions found at lower redshift, disfavoring strong evolution to z = 2.22. We estimate an amplification due to gravitational lensing of (1.10 0.23 mag) - compatible with the value estimated from the weak-lensing-derived mass and the mass-concentration relation from ΛCDM simulations - making it the most amplified SN Ia discovered behind a galaxy cluster.

We present a novel technique for fitting restframe I-band light curves on a data set of 42 type la Supernovae (SNe Ia). Using the result of the fit, we construct a Hubble diagram with 26 SNe from the subset at 0.01 < z < 0.1. Adding two SNe at z ∼ 0.5 yields results consistent with a flat Λ-dominated "concordance universe" (ΩM, ΩΛ) = (0.25, 0.75). For one of these, SN 2000fr, new near infrared data are presented. The high redshift supernova NIR data are also used to test for systematic effects in the use of SNe la as distance estimators. A flat, Λ = 0, universe where the faintness of supernovae at z ∼ 0.5 is due to grey dust homogeneously distributed in the intergalactic medium is disfavoured based on the high-z Hubble diagram using this small data-set. However, the uncertainties are large and no firm conclusion may be drawn. We explore the possibility of setting limits on intergalactic dust based on B - I and B - V colour measurements, and conclude that about 20 well measured SNe are needed to give statistically significant results. We also show that the high redshift restframe I-band data points are better fit by light curve templates that show a prominent second peak, suggesting that they are not intrinsically underluminous. © ESO 2005.

We present a weak-lensing study of SPT-CL J2040-4451 and IDCS J1426+3508 at z = 1.48 and 1.75, respectively. The two clusters were observed in our "See Change" program, a Hubble Space Telescope survey of 12 massive high-redshift clusters aimed at high-z supernova measurements and weak-lensing estimation of accurate cluster masses. We detect weak but significant galaxy shape distortions using infrared images from the Wide Field Camera 3 (WFC3), which has not yet been used for weak-lensing studies. Both clusters appear to possess relaxed morphology in projected mass distribution, and their mass centroids agree nicely with those defined by both the galaxy luminosity and X-ray emission. Using a Navarro-Frenk-White profile, for which we assume that the mass is tightly correlated with the concentration parameter, we determine the masses of SPT-CL J2040-4451 and IDCS J1426 + 3508 to be M200 = 8.6+1.7-1.4 × 1014 Moand 2.2+1.1-0.7 × 1014Mo, respectively. The weak-lensing mass of SPT-CL J2040-4451 shows that the cluster is clearly a rare object. Adopting the central value, the expected abundance of such a massive cluster at z ≳1.48 is only ∼ 0.07 in the parent 2500 sq. deg. survey. However, it is yet premature to claim that the presence of this cluster creates a serious tension with the current ΛCDM paradigm unless that tension will remain in future studies after marginalizing over many sources of uncertainties such as the accuracy of the mass function and the mass-concentration relation at the high-mass end. The mass of IDCS J1426+3508 is in excellent agreement with our previous Advanced Camera for Surveys-based weak-lensing result, while the much higher source density from our WFC3 imaging data makes the current statistical uncertainty ∼40% smaller.

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Telomere length, a reliable predictor of disease pathogenesis, can be affected by genetics, chronic stress and health behaviors. Cross-sectionally, highly stressed postmenopausal women have shorter telomeres, but only if they are inactive. However, no studies have prospectively examined telomere length change over a short period, and if rate of attrition is affected by naturalistic factors such as stress and engagement in healthy behaviors, including diet, exercise, and sleep. Here we followed healthy women over 1 year to test if major stressors that occurred over the year predicted telomere shortening, and whether engaging in healthy behaviors during this period mitigates this effect. In 239 postmenopausal, non-smoking, disease-free women, accumulation of major life stressors across a 1-year period predicted telomere attrition over the same period-for every major life stressor that occurred during the year, there was a significantly greater decline in telomere length over the year of 35 bp (P<0.05). Yet, these effects were moderated by health behaviors (interaction B=0.19, P=0.04). Women who maintained relatively higher levels of health behaviors (1 s.d. above the mean) appeared to be protected when exposed to stress. This finding has implications for understanding malleability of telomere length, as well as expectations for possible intervention effects. This is the first study to identify predictors of telomere length change over the short period of a year.

Introduction

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Discussion

In the P388D1 macrophage-like cell line, phospholipase A2 activity and prostaglandin production are stimulated by platelet-activating factor (PAF) and bacterial lipopolysaccharide (LPS). We have investigated the role of Ins(1,4,5)P3 and Ca2+ in signal transduction of PAF-induced prostaglandin E2 (PGE2) formation in these cells. The role of Ca2+ in the activation mechanism was studied by fluorescence imaging of intracellular Ca2+ in individual adherent cells and by determining the PGE2 production in the same population of cells. This new approach enabled us to correlate directly events on the single-cell level with a physiologically relevant response of the cell population. Priming the cells with LPS was required for PAF to stimulate PGE2 formation, yet LPS affected neither the intracellular free Ca2+ concentration ([Ca2+]i) nor the PAF-induced rise in [Ca2+]i. In addition, basal and PAF-stimulated Ins(1,4,5)P3 levels were not affected by LPS priming. However, the Ca2+ transient, the release of Ins(1,4,5)P3 and the formation of PGE2 induced by PAF were inhibited in cells pretreated with pertussis toxin. Buffering the [Ca2+]i with intracellular BAPTA [bis-(o-aminophenoxy)ethane-NNN'N'-tetra-acetic acid] blocked the PAF-stimulated rise in [Ca2+]i and PGE2 formation. Removal of extracellular Ca2+ during PAF stimulation prevented the influx of Ca2+, but did not affect the initial [Ca2+]i transient, nor did it inhibit PGE2 formation. Under the same conditions, ionomycin stimulated an identical [Ca2+]i transient, but, in contrast with PAF stimulation, no PGE2 formation was observed. PGE2 production could be rescued by prompt subsequent addition of PAF, which caused no further [Ca2+]i change on its own. These results show that the transient initial rise in [Ca2+]i, produced either by PAF via the formation of Ins(1,4,5)P3 or directly by ionomycin, is necessary, but not sufficient for the formation of PGE2 in LPS-primed P388D1 cells. Furthermore, we have demonstrated for the first time that PAF triggers a second signal that is not mediated by a change in [Ca2+]i. However, both signals are required to induce PGE2 formation.

We combine a novel boronate trap for F(-) with a near-infrared fluorophore into a single molecule. Attachment to targeting ligands enables localization by positron emission tomography (PET) and near-infrared fluorescence (NIRF). Our first application of this generic tag is to label Lymphoseek (tilmanocept), an agent designed for receptor-specific sentinel lymph node (SLN) mapping. The new conjugate incorporates (18)F(-) in a single, aqueous step, targets mouse SLN rapidly (1 h) with reduced distal lymph node accumulation, permits PET or scintigraphic imaging of SLN, and enables NIRF-guided excision and histological verification even after (18)F decay. This embodiment is superior to current SLN mapping agents such as nontargeted [(99m)Tc]sulfur colloids and Isosulfan Blue, as well as the phase III targeted ligand [(99m)Tc]SPECT Lymphoseek counterpart, species that are visible by SPECT or visible absorbance separately. Facile incorporation of (18)F into a NIRF probe should promote many synergistic PET and NIRF combinations.

A variety of genetically encoded reporters use changes in fluorescence (or Förster) resonance energy transfer (FRET) to report on biochemical processes in living cells. The standard genetically encoded FRET pair consists of CFPs and YFPs, but many CFP-YFP reporters suffer from low FRET dynamic range, phototoxicity from the CFP excitation light and complex photokinetic events such as reversible photobleaching and photoconversion. We engineered two fluorescent proteins, Clover and mRuby2, which are the brightest green and red fluorescent proteins to date and have the highest Förster radius of any ratiometric FRET pair yet described. Replacement of CFP and YFP with these two proteins in reporters of kinase activity, small GTPase activity and transmembrane voltage significantly improves photostability, FRET dynamic range and emission ratio changes. These improvements enhance detection of transient biochemical events such as neuronal action-potential firing and RhoA activation in growth cones.

Tungsten NAr^R alkylidene complexes have been prepared that contain the electron-withdrawing Ar^R groups 2,4,6-X₃C₆H₂ (Ar^X3, X = Cl, Br), 2,6-Cl₂-4-CF₃C₆H₂ (Ar^Cl2CF3), and 3,5-(CF₃)₂C₆H₃ (Ar^(CF3)2). Reported complexes include W(NAr^R)₂Cl₂(dme) (dme = 1,2-dimethoxyethane), W(NAr^R)₂(CH₂CMe₃)₂, W(NAr^R)(CHCMe₃)(OTf)₂(dme), and W(NAr^R)(CHCMe₃)(ODBMP)₂ (DBMP = 4-Me-2,6-(CHPh₂)C₆H₂). The W(NAr^R)(CHCMe₃)(ODBMP)₂ complexes were explored as initiators for the polymerization of 2,3-dicarbomethoxynorbornadiene (DCMNBD).

Nitriles are found in many bioactive compounds, and are among the most versatile functional groups in organic chemistry. Despite many notable recent advances, however, there are no approaches that may be used for the preparation of di- or tri-substituted alkenyl nitriles. Related approaches that are broad in scope and can deliver the desired products in high stereoisomeric purity are especially scarce. Here, we describe the development of several efficient catalytic cross-metathesis strategies, which provide direct access to a considerable range of Z- or E-di-substituted cyano-substituted alkenes or their corresponding tri-substituted variants. Depending on the reaction type, a molybdenum-based monoaryloxide pyrrolide or chloride (MAC) complex may be the optimal choice. The utility of the approach, enhanced by an easy to apply protocol for utilization of substrates bearing an alcohol or a carboxylic acid moiety, is highlighted in the context of applications to the synthesis of biologically active compounds.

Cis,syndiotacticA-alt-B copolymers, where A and B are two enantiomerically pure trans-2,3-disubstituted-5,6-norbornenes with "opposite" chiralities, can be prepared with stereogenic-at-metal initiators of the type M(NR)(CHR')(OR")(pyrrolide). Formation of a high percentage of alternating AB copolymer linkages relies on an inversion of chirality at the metal with each propagating step and a relatively fast formation of an AB sequence as a consequence of a preferred diastereomeric relationship between the chirality at the metal and the chirality of the monomer. This approach to formation of an alternating AB copolymer contrasts dramatically with the principle of forming AB copolymers from achiral monomers and catalysts.

Over the U.S. business cycle, fluctuations in residential investment are well known to systematically lead GDP. These dynamics are documented here to be specific to the U.S. and Canada. In other developed economies residential investment is broadly coincident with GDP. Nonresidential investment has the opposite dynamics, being coincident with or lagging GDP. These observations are in sharp contrast with the properties of nearly all business cycle models with disaggregated investment. Including mortgages and interest rate dynamics aligns the theory more closely with U.S. observations. Longer time to build in housing construction makes residential investment coincident with output.