The centrosomal Deubiquitylase USP21 regulates Gli1 transcriptional activity and stability

USP21 is a centrosome-associated deubiquitylase (DUB) that has been implicated in the formation of primary cilia – crucial organelles for the regulation of the Hedgehog (Hh) signaling pathway in vertebrates. Here, we identify KCTD6 – a cullin-3 E3-ligase substrate adapter that has been previously linked to Hh signaling – as well as Gli1, the key transcription factor responsible for Hh signal amplification, as new interacting partners of USP21. We identify a cryptic structured protein interaction domain in KCTD6, which is predicted to have a similar fold to Smr domains. Importantly, we show that both depletion and overexpression of catalytically active USP21 suppress Gli1-dependent transcription. Gli proteins are negatively regulated through protein kinase A (PKA)-dependent phosphorylation. We provide evidence that USP21 recruits and stabilises Gli1 at the centrosome where it promotes its phosphorylation by PKA. By revealing an intriguing functional pairing between a spatially restricted deubiquitylase and a kinase, our study highlights the centrosome as an important hub for signal coordination

Mildred Dresselhaus and Solid State Pedagogy at MIT

Mildred Dresselhaus is known for her influential research on the physics of carbon. Her wide‐ranging influence as a physics teacher, although well‐known to her students, has been less thoroughly examined. Exploring how Dresselhaus grew into her role teaching solid state physics at MIT reveals much about how that subfield evolved

Shock compression of molten silicate: results for a model basaltic composition

A technique has been developed for measurement of the shock wave, pressure-density equation of state of molten silicates initially at temperatures of up to ∼2000 K. A 40-mm propellant gun apparatus accelerates metal flyer plates to speeds of up to 2.5 km s^(−1); these flyer plates are capable of driving shock waves with amplitudes of 35–40 GPa (350–400 kbar) into molten silicate samples. Modifications to the standard equation of state experiments that are described here include design of a molybdenum sample container for the molten silicate; use of a 10-kW radio frequency induction heater to melt the sample prior to impact; implementation of shuttering systems to protect the optical system and prevent preexposure of the film in the rotating-mirror, continuously writing, streak camera; and reduction of Hugoniot data taking into account the effect of the sample capsule. Data for a model basaltic liquid (36 mol % anorthite, 64 mol % diopside) at an initial temperature of 1673 K and initial density of 2.61 Mg m^(−3), yield a shock velocity-particle velocity (U_(S)-U_(P)) relation given by U_S = 3.06 + 1.36 UP km s^(−1) up to values of U_P = 1.7 km s^(−1). The zero-pressure, bulk sound speed is in good agreement with ultrasonic measurements. The best fit Birch-Murnaghan equation of state for this model basaltic liquid is K_(0S) = 24.2 GPa and K′ = 4.85 based on Hugoniot points at low pressures (<25 GPa). Within the resolution of our data set, density increases smoothly with pressure over the 0–25 GPa pressure range, suggesting that structural rearrangements take place gradually in response to pressure in this pressure interval. At high pressures (≳ 25 GPa) the Hugoniot data suggest that the liquid stiffens considerably. This may indicate that the gradual structural changes characteristic of the lower-pressure regime, such as changes of Al3+ and Si4+ coordination by oxygen from fourfold to sixfold, are essentially complete by ∼25 GPa. These high-pressure Hugoniot data are fit by US = 0.85 + 2.63 Up km s^(−1). The high-pressure regime is similar to that obtained in initially solid silicates upon shock compression. Shock temperature calculations yield values of 2400–2600 K at 25 GPa, and the states achieved are believed to lie metastably in the liquid field

Plasmodium falciparum: linkage disequilibrium between loci in chromosomes 7 and 5 and chloroquine selective pressure in Northern Nigeria.

In view of the recent discovery (Molecular Cell 6, 861-871) of a (Lys76Thr) codon change in gene pfcrt on chromosome 7 which determines in vitro chloroquine resistance in Plasmodium falciparum, we have re-examined samples taken before treatment in our study in Zaria, Northern Nigeria (Parasitology, 119, 343-348). Drug resistance was present in 5/5 cases where the pfcrt 76Thr codon change was seen (100% positive predictive value). Drug sensitivity was found in 26/28 cases where the change was absent (93% negative predictive value). Allele pfcrt 76Thr showed strong linkage disequilibrium with pfmdr1 Tyr86 on chromosome 5, more complete than that between pfcrt and cg2 alleles situated between recombination cross-over points on chromosome 7. Physical linkage of cg2 with pfcrt may account for linkage disequilibrium between their alleles but in the case of genes pfmdr1 and pfcrt, on different chromosomes, it is likely that this is maintained epistatically through the selective pressure of chloroquine

The structure of the D49 phospholipase A(2) piratoxin III from Bothrops pirajai reveals unprecedented structural displacement of the calcium-binding loop: possible relationship to cooperative substrate binding

Snake venoms are rich sources of phospholipase A(2) homologues, both active calcium-binding Asp49 enzymes and essentially inactive Lys49 proteins. They are responsible for multiple pharmacological effects, some of which are dependent on catalytic activity and others of which are not. Here, the 2.4 Angstrom X-ray crystal structure of an active Asp49 phospholipase A(2) from the venom of the snake Bothrops pirajai, refined to conventional and free R values of 20.1 and 25.5%, respectively, is reported. Unusually for phospholipases A(2), the dependence of the enzyme rate on the substrate concentration is sigmoidal, implying cooperativity of substrate binding. The unprecedented structural distortion seen for the calcium-binding loop in the present structure may therefore be indicative of a T-state enzyme. An explanation of the interaction between the substrate-binding sites based on the canonical phospholipase A(2) dimer is difficult. However, an alternative putative dimer interface identified in the crystal lattice brings together the calcium-binding loops of neighbouring molecules, along with the C-terminal regions which are disulfide bonded to those loops, thereby offering a possible route of communication between active sites.59225526

The Structure Of The D49 Phospholipase A2 Piratoxin Iii From Bothrops Pirajai Reveals Unprecedented Structural Displacement Of The Calcium-binding Loop: Possible Relationship To Cooperative Substrate Binding

Snake venoms are rich sources of phospholipase A2 homologues, both active calcium-binding Asp49 enzymes and essentially inactive Lys49 proteins. They are responsible for multiple pharmacological effects, some of which are dependent on catalytic activity and others of which are not. Here, the 2.4 Å X-ray crystal structure of an active Asp49 phospholipase A2 from the venom of the snake Bothrops pirajai, refined to conventional and free R values of 20.1 and 25.5%, respectively, is reported. Unusually for phospholipases A2, the dependence of the enzyme rate on the substrate concentration is sigmoidal, implying cooperativity of substrate binding. The unprecedented structural distortion seen for the calcium-binding loop in the present structure may therefore be indicative of a T-state enzyme. An explanation of the interaction between the substrate-binding sites based on the canonical phospholipase A2 dimer is difficult. 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Shock wave apparatus for studying minerals at high pressure and impact phenomena on planetary surfaces

Shock wave and experimental impact phenomena research on geological and planetary materials is being carried out using two propellant (18 and 40 mm) guns (up to 2.5 km/sec) and a two‐stage light gas gun (up to 7 km/sec). Equation of state measurements on samples initially at room temperature and at low and high temperatures are being conducted using the 40 mm propellant apparatus in conjunction with Helmholtz coils, and radiative detectors and, in the case of the light gas gun, with streak cameras. The 18 mm propellant gun is used for recovery experiments on minerals, impact on cryogenic targets, and radiative post‐shock temperature measurements

Molecular modeling and inhibitory activity of cowpea cystatin against bean bruchid pests.

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Identification of Nucleases and Phosphatases by Direct Biochemical Screen of the Saccharomyces cerevisiae Proteome

The availability of yeast strain collections expressing individually tagged proteins to facilitate one-step purification provides a powerful approach to identify proteins with particular biochemical activities. To identify novel exo- and endo-nucleases that might function in DNA repair, we undertook a proteomic screen making use of the movable ORF (MORF) library of yeast expression plasmids. This library consists of 5,854 yeast strains each expressing a unique yeast ORF fused to a tripartite tag consisting of His6, an HA epitope, a protease 3C cleavage site, and the IgG-binding domain (ZZ) from protein A, under the control of the GAL1 promoter for inducible expression. Pools of proteins were partially purified on IgG sepharose and tested for nuclease activity using three different radiolabeled DNA substrates. Several known nucleases and phosphatases were identified, as well as two new members of the histidine phosphatase superfamily, which includes phosphoglycerate mutases and phosphatases. Subsequent characterization revealed YDR051c/Det1 to be an acid phosphatase with broad substrate specificity, whereas YOR283w has a broad pH range and hydrolyzes hydrophilic phosphorylated substrates. Although no new nuclease activities were identified from this screen, we did find phosphatase activity associated with a protein of unknown function, YOR283w, and with the recently characterized protein Det1. This knowledge should guide further genetic and biochemical characterization of these proteins