Structure-based Design and Functional studies of Novel Noroviral 3C Protease Chimeras Offers Insights into Substrate Specificity
This study also shows that chimeric murine noroviruses provide improved models for studying human norovirus protein function in the context of a full ORF1 polyprotein. (Source: BJ Cell)
Source: BJ Cell - October 2, 2014 Category: Biochemistry Authors: M R. Herod, C A. Prince, R J. Skilton, V K. Ward, J B. Cooper, I N. Clarke Tags: BJ Disease Source Type: research
Inpp5e increases the Rab5 association and phosphatidylinositol 3-phosphate accumulation at the phagosome through an interaction with Rab20
Phosphoinositide 5-phosphatases have been implicated in the regulation of phagocytosis. However, their precise roles in the phagocytic process are poorly understood. We prepared RAW264.7 macrophages deficient in Inpp5e (shInpp5e) to clarify the role of this lipid phosphatase. In shInpp5e cells, the uptake of solid particles was increased, and the rate of phagosome acidification was accelerated. As expected, levels of PtdIns(3,4,5)P3 and PtdIns(3,4)P2 were increased and decreased, respectively, on the forming phagocytic cups of these cells. Unexpectedly, the most prominent consequence of the Inpp5e deficiency was the decrea...
Source: BJ Cell - October 1, 2014 Category: Biochemistry Authors: T Segawa, K Hazeki, K Nigorikawa, S Morioka, Y Guo, S Takasuga, K Asanuma, O Hazeki Tags: BJ Cell Source Type: research
A protein kinase screen of Neurospora crassa mutant strains reveals that the SNF1 protein kinase promotes glycogen synthase phosphorylation
Glycogen functions as a carbohydrate reserve in a variety of organisms and its metabolism is highly regulated. The activities of glycogen synthase and glycogen phosphorylase, the rate-limiting enzymes of the synthesis and degradation processes, respectively, are regulated by allosteric modulation and by reversible phosphorylation. To identify the protein kinases affecting glycogen metabolism in N. crassa, we performed a screen of 84 serine/threonine kinase knockout strains. We identified multiple kinases that have already been described as controlling glycogen metabolism in different organisms, such as NcSNF1, NcPHO85, NcG...
Source: BJ Cell - September 25, 2014 Category: Biochemistry Authors: T De Souza Candido Jr., R Duarte Gonçalves, A Felicio, F Zanolli Freitas, F Barbosa Cupertino, A Gomes Vieira De Carvalho, M Bertolini Sr. Tags: BJ Biomolecules Source Type: research
A context-independent N-glycan signal targets the misfolded extracellular domain of Arabidopsis STRUBBELIG to endoplasmic reticulum-associated degradation
N-glycosylation of proteins plays an important role for the determination of the fate of newly synthesized glycoproteins in the endoplasmic reticulum (ER). Specific oligosaccharide structures recruit molecular chaperones that promote folding or mannose binding lectins that assist in clearance of improperly folded glycoproteins by delivery to ER-associated degradation (ERAD). In plants, the mechanisms and factors that recognize non-native proteins and sort them to ERAD are poorly understood. Here, we provide evidence that a misfolded variant of the STRUBBELIG extracellular domain (SUBEX-C57Y) is degraded in a glycan-depende...
Source: BJ Cell - September 24, 2014 Category: Biochemistry Authors: S Hüttner, C Veit, U Vavra, J Schoberer, M Dicker, D Maresch, F Altmann, R Strasser Tags: BJ Plant Source Type: research
Identification of Amino Acid Determinants in CYP4B1 for Optimal Catalytic Processing of 4-Ipomeanol
Mammalian CYP4B1 enzymes are cytochrome P450 monooxygenases that are responsible for the bioactivation of several exogenous pro-toxins including 4-ipomeanol (4-IPO). In contrast to the orthologous rabbit enzyme, we show here that native human CYP4B1 with a serine at position 427 is unable to bio-activate 4-IPO and does not cause cytotoxicity in HepG2 cells and primary human T-cells that overexpress these enzymes. We also demonstrate that a proline residue in the meander region at position 427 in human and 422 in the rabbit CYP4B1 is important for protein stability and rescues the 4-IPO bioactivation of the human enzyme, bu...
Source: BJ Cell - September 23, 2014 Category: Biochemistry Authors: C Wiek, E Maria Schmidt, K Röllecke, M Freund, M Nakano, E J Kelly, W Kaisers, V Yarov-Yarovoy, C M Kramm, A E Rettie, H Hanenberg Tags: BJ Cell Source Type: research
Sulphonylurea receptors regulate the channel pore in ATP-sensitive potassium channels via an inter-subunit salt bridge
ATP-sensitive potassium channels play key roles in many tissues by coupling metabolic status to membrane potential. In contrast to other potassium channels, the pore-forming Kir6 subunits must co-assemble in hetero-octameric complexes with ABC family sulphonylurea receptor (SUR) subunits to facilitate cell surface expression. Binding of nucleotides and drugs to SUR regulates channel gating but how these responses are communicated within the complex, has remained elusive. We have now identified an electrostatic interaction, forming part of a functional interface between the cytoplasmic nucleotide binding domain-2 of SUR2 su...
Source: BJ Cell - September 19, 2014 Category: Biochemistry Authors: D Lodwick, R D Rainbow, H N Rubaiy, M Al Johi, G W Vuister, R I Norman Tags: BJ Energy Source Type: research
Arp2/3 complex regulates adipogenesis by controlling cortical actin remodeling
Extensive actin cytoskeleton remodeling occurs during adipocyte development. We previously showed that disruption of stress fibers by the actin-severing protein cofilin is a requisite step in adipogenesis. However, it remains unclear whether actin nucleation and assembly into the cortical structure is essential for adipocyte development. Here we investigated the role of cortical actin assembly and of actin nucleation by the Arp2/3 complex in adipogenesis. Cortical actin structure formation started with accumulation of F-actin patches near the plasma membrane during adipogenesis. Depletion of Arp2/3 by knockdown of its subu...
Source: BJ Cell - September 15, 2014 Category: Biochemistry Authors: W Yang, S Thein, C Lim, R E Ericksen, S Sugii, F Xu, R C. Robinson, J Kim, W Han Tags: BJ Cell Source Type: research
Doc2b Serves as a Scaffolding Platform for Concurrent Binding of Multiple Munc18 Isoforms in Pancreatic Islet Beta Cells
Biphasic glucose-stimulated insulin secretion (GSIS) from pancreatic beta cells involves SNARE protein-regulated exocytosis. SNARE complex assembly further requires the regulatory proteins Munc18c, Munc18-1 and Doc2b. Munc18-1 and Munc18c are required for 1st- and 2nd-phase GSIS, respectively. These distinct Munc18-1 and Munc18c roles are related to their transient high-affinity binding with their cognate t-SNAREs; Syntaxin 1A and Syntaxin 4, respectively. Doc2b is essential for both phases of GSIS, yet the molecular basis for this remains unresolved. Because Doc2b binds to Munc18-1 and Munc18c via it’s distinct C2A...
Source: BJ Cell - September 5, 2014 Category: Biochemistry Authors: L Ramalingam, J Lu, A Hudmon, D C. Thurmond Tags: BJ Metabolism Source Type: research
MEKK2 regulates paxillin ubiquitylation and localization in MDA-MB 231 breast cancer cells
In this study we investigate MEKK2 function in focal adhesions, and we report that MEKK2 physically associates with the LD1 motif of the focal adhesion protein paxillin. We reveal that MEKK2 induces paxillin ubiquitylation, and that this function requires both the paxillin’s LD1 motif and MEKK2 kinase activity. Finally, we demonstrate that MEKK2 promotes paxillin redistribution from focal adhesions into the cytoplasm, but does not promote paxillin degradation. Taken together, our results reveal a novel function for MEKK2 as a regulator of ubiquitylation-dependent paxillin redistribution in breast tumor cells. (Source: BJ Cell)
Source: BJ Cell - September 5, 2014 Category: Biochemistry Authors: M Ameka, M P. Kahle, M Perez-Neut, S Gentile, A A. Mirza, B D Cuevas Tags: BJ Signal Source Type: research
The antimicrobial peptide LL-37 facilitates the formation of neutrophil extracellular traps
In conclusion, this study demonstrates a novel role of LL-37 in the formation of NETs. (Source: BJ Cell)
Source: BJ Cell - September 2, 2014 Category: Biochemistry Authors: A Neumann, E Berends, A Nerlich, E Molhoek, R Gallo, T Meerloo, V Nizet, H Y Naim, M von Köckritz-Blickwede Tags: BJ Cell Source Type: research
Molecular and structural transmembrane determinants critical for embedding claudin-5 into tight junctions reveal distinct four helix bundle arrangement
The mechanism of tight junction (TJ) assembly and the structure of TJ strand-forming claudins (Cldn) are unclear. To identify determinants of assembly of blood brain barrier-related Cldn3 and Cldn5, chimeric mutants were analyzed by cellular reconstitution of TJ strands and live cell imaging. Based on rescue of mutants deficient for strand formation, we identified Cldn5 residues involved in claudin folding (A132, A163, I166, L174) and assembly (C128, I142). Experimental results were combined with structural bioinformatics approaches. Initially the experimentally validated previous model of the extracellular loop 2 (ECL2) o...
Source: BJ Cell - September 1, 2014 Category: Biochemistry Authors: J Rossa, J Protze, C Kern, A Piontek, D Günzel, G Krause, J Piontek Tags: BJ Biomolecules Source Type: research
Proton-associated sucrose transport of mammalian Solute Carrier Family 45: an analysis in Saccharomyces cerevisiae
The members of the solute carrier family 45 (SLC45) have been implicated with the regulation of glucose homeostasis in the brain (SLC45A1), with skin and hair pigmentation (SLC45A2), and with prostate cancer and myelination (SLC45A3). However, apart from A1, a proton-associated glucose transporter, the function of these proteins is still largely unknown although sequence similarities to plant sucrose transporters mark them as a putative sucrose transporter family. Heterologous expression of the three members SLC45A2, SLC45A3 and SLC45A4 in Saccharomyces cerevisiae confirmed that they are indeed sucrose transporters. 14C-su...
Source: BJ Cell - August 28, 2014 Category: Biochemistry Authors: R Bartölke, J J Heinisch, H Wieczorek, O Vitavska Tags: BJ Cell Source Type: research
Red fluorescent genetically encoded Ca2{+} indicators for use in mitochondria and endoplasmic reticulum
Ca2+ is a key intermediary in a variety of signaling pathways and undergoes dynamic changes in its cytoplasmic concentration due to release from stores within the endoplasmic reticulum (ER) and influx from the extracellular environment. In addition to regulating cytoplasmic Ca2+ signals, these responses also affect the [Ca2+] within the ER and mitochondria. Single fluorescent protein -based Ca2+ indicators, such as the GCaMP series based on the green fluorescent protein (GFP), are powerful tools for imaging changes in the concentration of Ca2+ associated with intracellular signaling pathw...
Source: BJ Cell - August 28, 2014 Category: Biochemistry Authors: J Wu, D Lawrence Prole, Y Shen, Z Lin, A Gnanasekaran, Y Liu, L Chen, H Zhou, S Chen, Y M Usachev, C W. Taylor, R E Campbell Tags: BJ ChemBio Source Type: research
Testosterone increases renal anti-aging klotho gene expression via the androgen receptor-mediated pathway
Gender is known to be associated with longevity and estrogen administration induced longevity-associated gene expressions is one of the potential mechanisms underlying the benefits of estrogen on life span, whereas the role of testosterone in the regulation of longevity-associated gene expressions remains largely unclear. The klotho gene, predominantly expressed in the kidney, has recently been discovered to be an aging suppressor gene. Here, we investigated the regulatory effects of testosterone on renal klotho gene expression in vivo and in vitro. In testosterone-administered mouse kidney and NRK-52E cells, increased klo...
Source: BJ Cell - August 27, 2014 Category: Biochemistry Authors: S Hsu, S Huang, S Lin, S Ka, A Chen, M Shih, Y Hsu Tags: BJ Gene Source Type: research
SIRT3 interactions with FOXO3 acetylation, phosphorylation and ubiquitinylation mediate endothelial cell responses to hypoxia
The endothelial cells (ECs) that line the vascular lumen are exposed to a wide variety of environmental stresses, such as hypoxia. Maladaptation to stress in ECs is a key event in the development of cardiovascular disease. Sirtuin-3 (SIRT3) is an NAD+-dependent protein deacetylase that modulates various proteins to control mitochondrial function and metabolism. We found that hypoxia elicits an increase in SIRT3 mRNA and protein expression in ECs. Under the same hypoxic conditions, the forkhead box class O transcription factor FOXO3 is deacetylated by SIRT3. The SIRT3-mediated deacetylation of FOXO3 further reduces F...
Source: BJ Cell - August 27, 2014 Category: Biochemistry Authors: A H.-H. Tseng, L Wu, S Shieh, D Ling Wang Tags: BJ Signal Source Type: research
