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https://www.readbyqxmd.com/read/28644030/a-plc-%C3%AE-1-feedback-pathway-regulates-lck-substrate-phosphorylation-at-the-t-cell-receptor-and-slp-76-complex
#1
Judson Belmont, Tao Gu, Ashley Marie Mudd, Arthur R Salomon
Phospholipase C gamma 1 (PLC-γ1) occupies a critically important position in the T cell signaling pathway. While its functions as a regulator of both Ca2+ signaling and PKC-family kinases are well characterized, PLC-γ1's role in the regulation of early T cell receptor signaling events is incompletely understood. Activation of the T cell receptor leads to the formation of a signalosome complex between SLP-76, LAT, PLC-γ1, Itk, and Vav1. Recent studies have revealed the existence of both positive and negative feedback pathways from SLP-76 to the apical kinase in the pathway, Lck...
June 23, 2017: Journal of Proteome Research
https://www.readbyqxmd.com/read/28638064/quantitative-phosphoproteomics-reveals-a-role-for-collapsin-response-mediator-protein-2-in-pdgf-induced-cell-migration
#2
Adil R Sarhan, Justyna Szyroka, Shabana Begum, Michael G Tomlinson, Neil A Hotchin, John K Heath, Debbie L Cunningham
The Platelet Derived Growth Factor (PDGF) family of ligands have well established functions in the induction of cell proliferation and migration during development, tissue homeostasis and interactions between tumours and stroma. However, the mechanisms by which these actions are executed are incompletely understood. Here we report a differential phosphoproteomics study, using a SILAC approach, of PDGF-stimulated mouse embryonic fibroblasts (MEFs). 116 phospho-sites were identified as up-regulated and 45 down-regulated in response to PDGF stimulation...
June 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28636172/a-phos-tag-based-micropipette-tip-method-for-rapid-and-selective-enrichment-of-phosphopeptides
#3
Elena Tianfei Yuan, Yoko Ino, Maho Kawaguchi, Yayoi Kimura, Hisashi Hirano, Emiko Kinoshita-Kikuta, Eiji Kinoshita, Tohru Koike
Phosphorylated peptides are attractive targets in the study of the phosphoproteome. Here, we introduce a simple and convenient micropipette-tip method for the separation of phosphorylated and nonphosphorylated peptides by using a phosphate-binding zinc(II) complex of 1,3-bis(pyridin-2-ylmethylamino)propan-2-olate (Phos-tag). A 200-μL micropipette tip containing 10 μL of swollen agarose beads functionalized with Phos-tag moieties was prepared. All steps in the phosphate-affinity separation (binding, washing, and elution) were conducted by using aqueous buffers at neutral pH values...
June 21, 2017: Electrophoresis
https://www.readbyqxmd.com/read/28634298/analyses-of-pde-regulated-phosphoproteomes-reveal-unique-and-specific-camp-signaling-modules-in-t-cells
#4
Michael-Claude G Beltejar, Ho-Tak Lau, Martin G Golkowski, Shao-En Ong, Joseph A Beavo
Specific functions for different cyclic nucleotide phosphodiesterases (PDEs) have not yet been identified in most cell types. Conventional approaches to study PDE function typically rely on measurements of global cAMP, general increases in cAMP-dependent protein kinase (PKA), or the activity of exchange protein activated by cAMP (EPAC). Although newer approaches using subcellularly targeted FRET reporter sensors have helped define more compartmentalized regulation of cAMP, PKA, and EPAC, they have limited ability to link this regulation to downstream effector molecules and biological functions...
June 20, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28634201/phosphoproteomic-profiling-reveals-alk-and-met-as-novel-actionable-targets-across-synovial-sarcoma-subtypes
#5
Emmy D G Fleuren, Myrella Vlenterie, Winette van der Graaf, Melissa H S Hillebrandt-Roeffen, James Blackburn, Xiuquan Ma, Howard Chan, Mandy C Magias, Anke van Erp, Laurens van Houdt, Sabri Cebeci, Amy van de Ven, Uta E Flucke, Erin E Heyer, David M Thomas, Christopher J Lord, Kieren D Marini, Vijesh Vaghjiani, Tim Mercer, Jason E Cain, Jianmin Wu, Yvonne M H Versleijen-Jonkers, Roger J Daly
Despite intensive multi-modal treatment of sarcomas, a heterogeneous group of malignant tumors arising from connective tissue, survival remains poor. Candidate-based targeted treatments have demonstrated limited clinical success, urging an unbiased and comprehensive analysis of oncogenic signaling networks to reveal therapeutic targets and personalized treatment strategies. Here we applied mass spectrometry-based phosphoproteomic profiling to the largest and most heterogeneous set of sarcoma cell lines characterized to date and identified novel tyrosine phosphorylation patterns, enhanced tyrosine kinases in specific subtypes, and potential driver kinases...
June 20, 2017: Cancer Research
https://www.readbyqxmd.com/read/28634120/highly-reproducible-improved-label-free-quantitative-analysis-of-cellular-phosphoproteome-by-optimization-of-lc-ms-ms-gradient-and-analytical-column-construction
#6
Nagib Ahsan, Judson Belmont, Zhuo Chen, James G Clifton, Arthur R Salomon
Expanding the sequencing depth of the peptides with a statistically significant quantitative change derived from a biological stimulation is critical. Here we demonstrate that optimization of LC gradient and analytical column construction can reveal over 30,000 unique peptides and 23,000 phosphopeptides at high confidence. The quantitative reproducibility of different analytical workflows was evaluated by comparing the phosphoproteome of CD3/4 stimulated and unstimulated T-cells as a model system. A fritless, 50cm-long column packed with 1...
June 17, 2017: Journal of Proteomics
https://www.readbyqxmd.com/read/28633965/phosphoproteome-analysis-reveals-a-critical-role-for-hedgehog-signalling-in-osteoblast-morphological-transitions
#7
Ariane Marumoto, Renato Milani, Rodrigo A da Silva, Célio Junior da Costa Fernandes, José Mauro Granjeiro, Carmen V Ferreira, Maikel P Peppelenbosch, Willian F Zambuzzi
The reciprocal and adaptive interactions between cells and substrates governing morphological transitions in the osteoblast compartment remain largely obscure. Here we show that osteoblast cultured in basement membrane matrix (Matrigel™) exhibits significant morphological changes after ten days of culture, and we decided to exploit this situation to investigate the molecular mechanisms responsible for guiding osteoblast morphological transitions. As almost all aspects of cellular physiology are under control of kinases, we generated more or less comprehensive cellular kinome profiles employing PepChip peptide arrays that contain over 1000 consensus substrates of kinase peptide...
June 17, 2017: Bone
https://www.readbyqxmd.com/read/28629377/quantitative-phosphoproteome-on-the-silkworm-bombyx-mori-cells-infected-with-baculovirus
#8
Jauharotus Shobahah, Shengjie Xue, Dongbing Hu, Cui Zhao, Ming Wei, Yanping Quan, Wei Yu
BACKGROUND: Bombyx mori has become an important model organism for many fundamental studies. Bombyx mori nucleopolyhedrovirus (BmNPV) is a significant pathogen to Bombyx mori, yet also an efficient vector for recombinant protein production. A previous study indicated that acetylation plays many vital roles in several cellular processes of Bombyx mori while global phosphorylation pattern upon BmNPV infection remains elusive. METHOD: Employing tandem mass tag (TMT) labeling and phosphorylation affinity enrichment followed by high-resolution LC-MS/MS analysis and intensive bioinformatics analysis, the quantitative phosphoproteome in Bombyx mori cells infected by BmNPV at 24 hpi with an MOI of 10 was extensively examined...
June 19, 2017: Virology Journal
https://www.readbyqxmd.com/read/28615362/drugging-the-catalytically-inactive-state-of-ret-kinase-in-ret-rearranged-tumors
#9
Dennis Plenker, Maximilian Riedel, Johannes Brägelmann, Marcel A Dammert, Rakhee Chauhan, Phillip P Knowles, Carina Lorenz, Marina Keul, Mike Bührmann, Oliver Pagel, Verena Tischler, Andreas H Scheel, Daniel Schütte, Yanrui Song, Justina Stark, Florian Mrugalla, Yannic Alber, André Richters, Julian Engel, Frauke Leenders, Johannes M Heuckmann, Jürgen Wolf, Joachim Diebold, Georg Pall, Martin Peifer, Maarten Aerts, Kris Gevaert, René P Zahedi, Reinhard Buettner, Kevan M Shokat, Neil Q McDonald, Stefan M Kast, Oliver Gautschi, Roman K Thomas, Martin L Sos
Oncogenic fusion events have been identified in a broad range of tumors. Among them, RET rearrangements represent distinct and potentially druggable targets that are recurrently found in lung adenocarcinomas. We provide further evidence that current anti-RET drugs may not be potent enough to induce durable responses in such tumors. We report that potent inhibitors, such as AD80 or ponatinib, that stably bind in the DFG-out conformation of RET may overcome these limitations and selectively kill RET-rearranged tumors...
June 14, 2017: Science Translational Medicine
https://www.readbyqxmd.com/read/28609623/single-step-enrichment-of-n-glycopeptides-and-phosphopeptides-with-novel-multifunctional-ti4-immobilized-dendritic-polyglycerol-coated-chitosan-nanomaterials
#10
Xiajuan Zou, Jianzheng Jie, Bin Yang
Protein glycosylation and phosphorylation, two of the most important post-translational modifications (PTMs) in the proteome play a vital role in regulating a number of complex biological processes and involvement in a variety of diseases. Comprehensive characterization of the phosphoproteome and glycoproteome requires highly specific and sensitive enrichment methods for purification of phosphopeptides and glycopeptides because many glycoproteins and phosphoproteins naturally occur at low abundances and substoichiometry...
June 13, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28600813/elucidation-of-altered-pathways-in-tumor-initiating-cells-of-triple-negative-breast-cancer-a-useful-cell-model-system-for-drug-screening
#11
Anne G Christensen, Sidse Ehmsen, Mikkel G Terp, Richa Batra, Nicolas Alcaraz, Jan Baumbach, Julie B Noer, José Moreira, Rikke Leth-Larsen, Martin R Larsen, Henrik J Ditzel
A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics...
June 10, 2017: Stem Cells
https://www.readbyqxmd.com/read/28588593/salinity-induced-palmella-formation-mechanism-in-halotolerant-algae-dunaliella-salina-revealed-by-quantitative-proteomics-and-phosphoproteomics
#12
Sijia Wei, Yangyang Bian, Qi Zhao, Sixue Chen, Jiawei Mao, Chunxia Song, Kai Cheng, Zhen Xiao, Chuanfang Zhang, Weimin Ma, Hanfa Zou, Mingliang Ye, Shaojun Dai
Palmella stage is critical for some unicellular algae to survive in extreme environments. The halotolerant algae Dunaliella salina is a good single-cell model for studying plant adaptation to high salinity. To investigate the molecular adaptation mechanism in salinity shock-induced palmella formation, we performed a comprehensive physiological, proteomics and phosphoproteomics study upon palmella formation of D. salina using dimethyl labeling and Ti(4+)-immobilized metal ion affinity chromatography (IMAC) proteomic approaches...
2017: Frontiers in Plant Science
https://www.readbyqxmd.com/read/28582757/comparative-analysis-of-muscle-phosphoproteome-induced-by-salt-curing
#13
Zhenyu Wang, Caixia Zhang, Zheng Li, Qingwu Shen, Dequan Zhang
Phosphorylated proteins in ovine muscle induced by salt curing were well investigated. Ten topside muscles of crossbred sheep were ground, mixed and divided into two groups, which were cured for 16h with 0% and 3% NaCl, respectively. Muscle proteins of two cured groups were analyzed by two-dimensional electrophoresis coupled with Pro-Q Diamond and SYPRO Ruby staining. The differential phosphorylated proteins were determined using LC-MS/MS and the UniProt database. Ten different phosphoproteins (>1.5 fold) induced by salting were identified, including triosephosphate isomerase, glycogen phosphorylase, creatine kinase M-type, myoglobin, troponin T fast skeletal muscle type, actin, myosin light chain 1/3, tropomyosin beta chain, etc...
May 23, 2017: Meat Science
https://www.readbyqxmd.com/read/28576409/phosphoproteome-and-transcription-factor-activity-profiling-identify-actions-of-the-anti-inflammatory-agent-utl-5g-in-lps-stimulated-raw-264-7-cells-including-disrupting-actin-remodeling-and-stat-3-activation
#14
Nicholas J Carruthers, Paul M Stemmer, Ben Chen, Frederick Valeriote, Xiaohua Gao, Subash C Guatam, Jiajiu Shaw
UTL-5g is a novel small-molecule TNF-alpha modulator. It reduces cisplatin-induced side effects by protecting kidney, liver, and platelets, thereby increasing tolerance for cisplatin. UTL-5g also reduces radiation-induced acute liver toxicity. The mechanism of action for UTL-5g is not clear at the present time. A phosphoproteomic analysis to a depth of 4943 phosphopeptides and a luminescence-based transcription factor activity assay were used to provide complementary analyses of signaling events that were disrupted by UTL-5g in RAW 264...
May 30, 2017: European Journal of Pharmacology
https://www.readbyqxmd.com/read/28574228/apkc-%C3%AE-p-sp1-snail-signaling-induces-epithelial-mesenchymal-transition-and-immunosuppression-in-cholangiocarcinoma
#15
Yawei Qian, Wei Yao, Tao Yang, Yan Yang, Yan Liu, Qi Shen, Jian Zhang, Weipeng Qi, Jianming Wang
Cholangiocarcinoma (CCA) is a highly malignant bile duct cancer that tends to invade and metastasize early. The epithelial-mesenchymal transition (EMT) has been implicated in cancer cell invasion and metastasis, as well as in cancer cell evasion of host immunity. In this study, we investigated the interaction between atypical protein kinase C-iota (aPKC-ι) and Snail in the regulation of EMT and its relationship to CCA immunosuppression. Our results demonstrated that aPKC-ι, Snail, and infiltrated immunosuppressive cells were significantly up-regulated in CCA tumor tissues and were linked to poor prognosis...
June 2, 2017: Hepatology: Official Journal of the American Association for the Study of Liver Diseases
https://www.readbyqxmd.com/read/28572185/stable-isotope-labeling-by-amino-acids-in-cell-culture-silac-based-quantitative-proteomics-and-phosphoproteomics-in-fission-yeast
#16
Alejandro Carpy, André Koch, Claudia C Bicho, Weronika E Borek, Silke Hauf, Kenneth E Sawin, Boris Maček
Modern mass spectrometry (MS)-based approaches are capable of identifying and quantifying thousands of proteins and phosphorylation events in a single biological experiment. Here we present a (phospho)proteomic workflow based on in-solution proteome digestion of samples labeled by stable isotope labeling by amino acids in cell culture (SILAC) and phosphopeptide enrichment using strong cation exchange (SCX) and TiO2 chromatographies. These procedures are followed by high-accuracy MS measurement on an Orbitrap mass spectrometer and subsequent bioinformatic processing using MaxQuant software...
June 1, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28556455/the-serine-threonine-phosphatases-of-apicomplexan-parasites
#17
REVIEW
Chunlin Yang, Gustavo Arrizabalaga
The balance between phosphorylation and de-phosphorylation, which is delicately regulated by protein kinases and phosphatases, is critical for nearly all biological processes. The Apicomplexa are a large phylum which contains various parasitic protists, including human pathogens, such as Plasmodium, Toxoplasma, Cryptosporidium and Babesia species. The diverse life cycles of these parasites are highly complex and, not surprisingly, many of their key steps are exquisitely regulated by phosphorylation. Interestingly, many of the kinases and phosphatases, as well as the substrates involved in these events are unique to the parasites and therefore phosphorylation constitutes a viable target for antiparasitic intervention...
May 27, 2017: Molecular Microbiology
https://www.readbyqxmd.com/read/28555341/offline-pentafluorophenyl-pfp-rp-prefractionation-as-an-alternative-to-high-ph-rp-for-comprehensive-lc-ms-ms-proteomics-and-phosphoproteomics
#18
Andrew V Grassetti, Rufus Hards, Scott A Gerber
Technological advances in liquid chromatography and tandem mass spectrometry (LC-MS/MS) have enabled comprehensive analyses of proteins and their post-translational modifications from cell culture and tissue samples. However, sample complexity necessitates offline prefractionation via a chromatographic method that is orthogonal to online reversed-phase high-performance liquid chromatography (RP-HPLC). This additional fractionation step improves target identification rates by reducing the complexity of the sample as it is introduced to the instrument...
May 29, 2017: Analytical and Bioanalytical Chemistry
https://www.readbyqxmd.com/read/28551142/simple-preparation-of-magnetic-metal-organic-frameworks-composite-as-a-bait-for-phosphoproteome-research
#19
Guobin Han, Qiaoling Zeng, Zhongwei Jiang, Wenchan Deng, Chengzhi Huang, Yuanfang Li
Phosphospecific enrichment techniques and mass spectrometry (MS) are primary tools for comprehending the cellular phosphoproteome. In this work, a rational and extremely facile route to synthesize the magnetic metal-organic frameworks (mMOFs) was employed and the prepared composite was first utilized as a "bait" for selective enrichment of phosphopeptides. Typically, the mMOFs was synthesized via electrostatic self-assembly between the negatively charged Fe3O4 magnetic nanoparticles (MNPs) and positively charged MIL-101(Fe)...
August 15, 2017: Talanta
https://www.readbyqxmd.com/read/28551015/salt-induced-phosphoproteomic-changes-in-the-hypothalamic-paraventricular-nucleus-in-rats-with-chronic-renal-failure
#20
Lanying Li, Jiawen Li, Lishan Tan, Minzi Qiu, Mengbi Zhang, Aiqing Li
Hypothalamic paraventricular nucleus (PVN) is a cardiovascular regulating center within the brain, which plays a critical role in high salt-induced progression of chronic renal failure (CRF). However, the phosphoproteomic changes in the PVN caused by CRF remain unclear. This study aimed to perform large-scale phosphoproteomic analysis of PVN induced by CRF and high salt intake. In this study, eight weeks post 5/6 nephrectomy (CRF model) or sham operation, Sprague-Dawley rats were fed a high-salt (4%) or normal-salt (0...
May 24, 2017: Brain Research
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