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Guttman-Yassky E, Brunner PM, Neumann AU, Khattri S, Pavel AB, Malik K, Singer GK, Baum D, Gilleaudeau P, Sullivan-Whalen M, Rose S, On SJ, Li X, Fuentes-Duculan J, Estrada Y, Garcet S, Traidl-Hoffmann C, Krueger JG, Lebwohl MG
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Efficacy and safety of fezakinumab (an IL-22 monoclonal antibody) in adults with moderate-to-severe atopic dermatitis inadequately controlled by conventional treatments: A randomized, double-blind, phase 2a trial

Background: Interleukin 22 promotes epidermal hyperplasia and inhibits skin barrier function. Objective: Evaluate interleukin 22 blockade in adults with moderate-to-severe atopic dermatitis (AD). Methods: We performed a randomized, double-blind, placebo-controlled trial with intravenous fezakinumab monotherapy every 2 weeks for 10 weeks, with follow-up assessments until 20 weeks. The change in SCOring AD (SCORAD) score from baseline at 12 weeks served as the primary end point. Results: At 12 weeks, the mean declines in SCORAD for the entire study population were 13.8 +/- 2.7 in the fezakinumab arm and 8.0 +/- 3.1 in the placebo arm (P=.134). In the severe AD patient subset (with a baseline SCORAD of >= 50), SCORAD decline was significantly stronger in the drug-treated patients than placebo-treated patients at 12 weeks (21.6 +/- 3.8 vs 9.6 +/- 4.2, P=.029) and 20 weeks (27.4 +/- 3.9 vs 11.5 +/- 5.1, P=.010). At 12 weeks, improvements in body surface area involvement in the entire population were significantly stronger in the drug-treated than placebo-treated patients (12.4% +/- 2.4 vs 6.2% +/- 2.7; P=.009), and in the severe AD subset, the decline in Investigator Global Assessment was significantly higher in the drug-treated than placebo-treated patients (0.7 +/- 0.2 vs 0.3 +/- 0.1; P=.034). All scores showed progressive improvements after last dosing (10 weeks) until end of study (20 weeks). Common adverse events were upper respiratory tract infections. Limitations: The limited sample size and lack of assessment with Eczema Area and Severity Index and a pruritus numerical rating scale were limiting factors. Significance was primarily obtained in severe AD. Conclusion: Fezakinumab was well-tolerated, with sustained clinical improvements after last drug dosing.
Irwin DM, Mojsov S
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Diversification of the functions of proglucagon and glucagon receptor genes in fish

The teleost fish-specific genome duplication gave rise to a great number of species inhabiting diverse environments with different access to nutrients and life histories. This event produced duplicated gcg genes, gcga and gcgb, for proglucagon-derived peptides, glucagon and GLP-1 and duplicated gcgr receptor genes, gcgra and gcgrb, which play key roles connecting the consumption of nutrients with glucose metabolism. We conducted a systematic survey of the genomes from 28 species of fish (24 bony (Superclass Osteichthyes), 1 lobe-finned (Class Sarcoperygii), 1 cartilaginous (Superclass Chondrichthyes), and 2 jaw less (Superclass Agnatha)) and find that almost all surveyed ray-finned fish contain gcga and gcgb genes with different coding potential and duplicated gcgr genes, gcgra and gcgrb that form two separate Glades in the phylogenetic tree consistent with the accepted species phylogeny. All gcgb genes encoded only glucagon and GLP-1 and gcga genes encoded glucagon, GLP-1, and GLP-2, indicating that gcga was subfunctionalized to produce GLP-2. We find a single glp2r, but no glp 1 r suggesting that duplicated gcgrb was neofunctionalized to bind GLP-1, as demonstrated for the zebrafish gcgrb (Oren et al., 2016). In functional experiments with zebrafish gcgrb and GLP-1 from diverse fish we find that anglerfish GLP-la, encoded by gcga, is less biologically active than the gcgb anglerfish GLP-1 b paralog. But some other fish (zebrafish, salmon, and catfish) gcga GLP-1 a display similar biological activities, indicating that the regulation of glucose metabolism by GLP-1 in ray-finned fish is species-specific. Searches of genomes in cartilaginous fish identified a proglucagon gene that encodes a novel GLP-3 peptide in addition to glucagon, GLP-1, and GLP-2, as well as a single gcgr, glp2r, and a new glucagon receptor-like receptor whose identity still needs to be confirmed. The sequence of the shark GLP-1 contained an N-terminal mammalian-like extension that in mammals undergoes a proteolytic cleavage to release biologically active GLP-1. Our results indicate that early in vertebrate evolution diverse regulatory mechanisms emerged for the control of glucose metabolism by proglucagon-derived peptides and their receptors and that in ray-finned fish they included subfunctionalization and neofunctionalization of these genes. (C) 2018 Elsevier Inc. All rights reserved.
Kodama H, Vroomen LG, Ueshima E, Reilly J, Brandt W, Paluch LR, Monette S, Jones D, Solomon SB, Srimathveeravalli G
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Catheter-based endobronchial electroporation is feasible for the focal treatment of peribronchial tumors

Objective: To evaluate the feasibility of catheter-based endobronchial electroporation for the treatment of peribronchial tumors and assess the incidence of treatment-related adverse events. Methods: Cytotoxicity of electroporation with or without cisplatin or gefitinib was assessed in vitro with lung cancer and normal cell lines. A novel catheter was designed for endobronchial electroporation, and computer simulations were used to predict in vivo treatment effects. Electroporation with the test catheter was performed (2000 V, 70 pulses) in the main bronchus of 8 pigs at 11 locations. Computed tomography imaging was performed before they were killed at 4 hours (6 animals) or 4 weeks (2 animals) posttreatment. Treated airway and surrounding parenchyma were compared with sham treatment via gross and histopathology. Results: Significant cell death due to electroporation and increased cytotoxicity in combination with cisplatin or gefitinib were observed in cancer cells only (P < .05). Simulations predicted penetrative electroporation of peribronchial parenchyma without tissue heating. Electric pulse delivery in vivo induced transient venous and bronchial spasms that resolved without intervention. Cross-sectional measurement of electroporation effects on computed tomography (14.4 +/- 1.4 by 10.5 +/- 1.3 mm) and gross pathology (17.2 +/- 3.0 by 8.8 +/- 0.6 mm) were representative of values predicted by simulation (P < .001). Cell death due to irreversible electroporation was observed in bronchial and parenchymal tissue in acute tissue samples. Treated lung rapidly recovered from the effects of electroporation without change in bronchial patency at 4 weeks posttreatment. Conclusions: Catheter-based endobronchial electroporation is a reproducible technique that can be used to treat peribronchial tumors in combination with cisplatin, without affecting patency of the treated bronchus.
Lee CH, MacKinnon R
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Activation mechanism of a human SK-calmodulin channel complex elucidated by cryo-EM structures

SCIENCE 2018 MAY 4; 360(6388):508-513
Small-conductance Ca2+-activated K+ (SK) channels mediate neuron excitability and are associated with synaptic transmission and plasticity. They also regulate immune responses and the size of blood cells. Activation of SK channels requires calmodulin (CaM), but how CaM binds and opens SK channels has been unclear. Here we report cryo-electron microscopy (cryo-EM) structures of a human SK4-CaM channel complex in closed and activated states at 3.4- and 3.5-angstrom resolution, respectively. Four CaM molecules bind to one channel tetramer. Each lobe of CaM serves a distinct function: The C-lobe binds to the channel constitutively, whereas the N-lobe interacts with the S4-S5 linker in a Ca2+-dependent manner. The S4-S5 linker, which contains two distinct helices, undergoes conformational changes upon CaM binding to open the channel pore. These structures reveal the gating mechanism of SK channels and provide a basis for understanding SK channel pharmacology.
Ge YJ, Fuchs E
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Stretching the limits: from homeostasis to stem cell plasticity in wound healing and cancer

Stem cells (SCs) govern tissue homeostasis and wound repair. They reside within niches, the special microenvironments within tissues that control SC lineage outputs. Upon injury or stress, new signals emanating from damaged tissue can divert nearby cells into adopting behaviours that are not part of their homeostatic repertoire. This behaviour, known as SC plasticity, typically resolves as wounds heal. However, in cancer, it can endure. Recent studies have yielded insights into the orchestrators of maintenance and lineage commitment for SCs belonging to three mammalian tissues: the haematopoietic system, the skin epithelium and the intestinal epithelium. We delineate the multifactorial determinants and general principles underlying the remarkable facets of SC plasticity, which lend promise for regenerative medicine and cancer therapeutics.
Geller R, Pechmann S, Acevedo A, Andino R, Frydman J
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Hsp90 shapes protein and RNA evolution to balance trade-offs between protein stability and aggregation

NATURE COMMUNICATIONS 2018 MAY 3; 9(?):? Article 1781
Acquisition of mutations is central to evolution; however, the detrimental effects of most mutations on protein folding and stability limit protein evolvability. Molecular chaperones, which suppress aggregation and facilitate polypeptide folding, may alleviate the effects of destabilizing mutations thus promoting sequence diversification. To illuminate how chaperones can influence protein evolution, we examined the effect of reduced activity of the chaperone Hsp90 on poliovirus evolution. We find that Hsp90 offsets evolutionary trade-offs between protein stability and aggregation. Lower chaperone levels favor variants of reduced hydrophobicity and protein aggregation propensity but at a cost to protein stability. Notably, reducing Hsp90 activity also promotes clusters of codon-deoptimized synonymous mutations at inter-domain boundaries, likely to facilitate cotranslational domain folding. Our results reveal how a chaperone can shape the sequence landscape at both the protein and RNA levels to harmonize competing constraints posed by protein stability, aggregation propensity, and translation rate on successful protein biogenesis.
Farber G, Hurtado R, Loh S, Monette S, Mtui J, Kopan R, Quaggin S, Meyer-Schwesinger C, Herzlinger D, Scott RP, Blobel CP
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Glomerular endothelial cell maturation depends on ADAM10, a key regulator of Notch signaling

ANGIOGENESIS 2018 MAY; 21(2):335-347
The principal function of glomeruli is to filter blood through a highly specialized filtration barrier consisting of a fenestrated endothelium, the glomerular basement membrane and podocyte foot processes. Previous studies have uncovered a crucial role of endothelial a disintegrin and metalloprotease 10 (ADAM10) and Notch signaling in the development of glomeruli, yet the resulting defects have not been further characterized nor understood in the context of kidney development. Here, we used several different experimental approaches to analyze the kidneys and glomeruli from mice lacking ADAM10 in endothelial cells (A10 Delta EC mice). Scanning electron microscopy of glomerular casts demonstrated enlarged vascular diameter and increased intussusceptive events in A10 Delta EC glomeruli compared to controls. Consistent with these findings, genes known to regulate vessel caliber (Apln, AplnR and Vegfr3) are significantly upregulated in A10 Delta EC glomeruli. Moreover, transmission electron microscopy revealed the persistence of diaphragms in the fenestrae of A10 Delta EC glomerular endothelial cells, which was corroborated by the elevated expression of the protein PLVAP/PV-1, an integral component of fenestral diaphragms. Analysis of gross renal vasculature by light sheet microscopy showed no major alteration of the branching pattern, indicating a localized importance of ADAM10 in the glomerular endothelium. Since intussusceptions and fenestrae with diaphragms are normally found in developing, but not mature glomeruli, our results provide the first evidence for a crucial role of endothelial ADAM10, a key regulator of Notch signaling, in promoting the development and maturation of the glomerular vasculature.
Blanco-Melo D, Gifford RJ, Bieniasz PD
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Reconstruction of a replication-competent ancestral murine endogenous retrovirus-L

RETROVIROLOGY 2018 MAY 2; 15(?):? Article 34
Background: About 10% of the mouse genome is composed of endogenous retroviruses (ERVs) that represent a molecular fossil record of past retroviral infections. One such retrovirus, murine ERV-L (MuERV-L) is an env-deficient ERV that has undergone episodic proliferation, with the most recent amplification occurring similar to 2 million years ago. MuERV-L related sequences have been co-opted by mice for antiretroviral defense, and possibly as promoters for some genes that regulate totipotency in early mouse embryos. However, MuERV-L sequences present in modern mouse genomes have not been observed to replicate. Results: Here, we describe the reconstruction of an ancestral MuERV-L (ancML) sequence through paleovirological analyses of MuERV-L elements in the modern mouse genome. The resulting MuERV-L (ancML) sequence was synthesized and a reporter gene embedded. The reconstructed MuERV-L (ancML) could replicate in a manner that is dependent on reverse transcription and generated de novo integrants. Notably, MuERV-L (ancML) exhibited a narrow host range. Interferon-a could reduce MuERV-L (ancML) replication, suggesting the existence of interferon-inducible genes that could inhibit MuERV-L replication. While mouse APOBEC3 was able to restrict the replication of MuERV-L (ancML), inspection of endogenous MuERV-L sequences suggested that the impact of APOBEC3 mediated hypermutation on MuERV-L has been minimal. Conclusion: The reconstruction of an ancestral MuERV-L sequence highlights the potential for the retroviral fossil record to illuminate ancient events and enable studies of the impact of retroviral elements on animal evolution.
Bieniasz PD, Kutluay SB
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CLIP-related methodologies and their application to retrovirology

RETROVIROLOGY 2018 MAY 2; 15(?):? Article 35
Virtually every step of HIV-1 replication and numerous cellular antiviral defense mechanisms are regulated by the binding of a viral or cellular RNA-binding protein (RBP) to distinct sequence or structural elements on HIV-1 RNAs. Until recently, these protein-RNA interactions were studied largely by in vitro binding assays complemented with genetics approaches. However, these methods are highly limited in the identification of the relevant targets of RBPs in physiologically relevant settings. Development of crosslinking-immunoprecipitation sequencing (CLIP) methodology has revolutionized the analysis of protein-nucleic acid complexes. CLIP combines immunoprecipitation of covalently crosslinked protein-RNA complexes with high-throughput sequencing, providing a global account of RNA sequences bound by a RBP of interest in cells (or virions) at near-nucleotide resolution. Numerous variants of the CLIP protocol have recently been developed, some with major improvements over the original. Herein, we briefly review these methodologies and give examples of how CLIP has been successfully applied to retrovirology research.
Mitchell MR, Leibler S
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Elastic strain and twist analysis of protein structural data and allostery of the transmembrane channel KcsA

PHYSICAL BIOLOGY 2018 MAY; 15(3):? Article 036004
The abundance of available static protein structural data makes the more effective analysis and interpretation of this data a valuable tool to supplement the experimental study of protein mechanics. Structural displacements can be difficult to analyze and interpret. Previously, we showed that strains provide a more natural and interpretable representation of protein deformations, revealing mechanical coupling between spatially distinct sites of allosteric proteins. Here, we demonstrate that other transformations of displacements yield additional insights. We calculate the divergence and curl of deformations of the transmembrane channel KcsA. Additionally, we introduce quantities analogous to bend, splay, and twist deformation energies of nematic liquid crystals. These transformations enable the decomposition of displacements into different modes of deformation, helping to characterize the type of deformation a protein undergoes. We apply these calculations to study the filter and gating regions of KcsA. We observe a continuous path of rotational deformations physically coupling these two regions, and, we propose, underlying the allosteric interaction between these regions. Bend, splay, and twist distinguish KcsA gate opening, filter opening, and filter-gate coupling, respectively. In general, physically meaningful representations of deformations (like strain, curl, bend, splay, and twist) can make testable predictions and yield insights into protein mechanics, augmenting experimental methods and more fully exploiting available structural data.