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phosphoproteomics rat

Niya Gowthami, B Sunitha, Manish Kumar, T S Keshava Prasad, N Gayathri, B Padmanabhan, M M Srinivas Bharath
In eukaryotes, mitochondrial complex I (NADH: ubiquinone oxidoreductase; CI) is central to oxidative phosphorylation (OXPHOS). Mammalian CI is a 45 subunit complex that forms supercomplexes with other OXPHOS complexes. Since CI defects are associated with aging and neurodegeneration, it is pertinent to understand its structure-function relationship. Although genetic mutations could lower CI activity causing mitochondrial dysfunction in several pathologies, post-translational modifications (PTMs) have emerged as a key mechanism contributing to altered CI activity...
February 27, 2018: Journal of Chemical Neuroanatomy
Gabriela Caraveo, Martin Soste, Valentina Cappelleti, Saranna Fanning, Damian B van Rossum, Luke Whitesell, Yanmei Huang, Chee Yeun Chung, Valeriya Baru, Sofia Zaichick, Paola Picotti, Susan Lindquist
Calcineurin is an essential Ca2+ -dependent phosphatase. Increased calcineurin activity is associated with α-synuclein (α-syn) toxicity, a protein implicated in Parkinson's Disease (PD) and other neurodegenerative diseases. Calcineurin can be inhibited with Tacrolimus through the recruitment and inhibition of the 12-kDa cis-trans proline isomerase FK506-binding protein (FKBP12). Whether calcineurin/FKBP12 represents a native physiologically relevant assembly that occurs in the absence of pharmacological perturbation has remained elusive...
December 26, 2017: Proceedings of the National Academy of Sciences of the United States of America
Zeng Li, Zhen Wang, Shun Xu, Wenwei Liang, Weimin Fan
BACKGROUND/AIMS: In recent years, a variety of studies have been performed to investigate the cellular responses of periodic mechanical stress. In our previous studies, we found that periodic mechanical stress can promote proliferation and matrix synthesis through the integrin beta 1-mediated ERK1/2 pathway, and we used proteomic analysis to detect quantitative changes in chondrocytes under periodic mechanical stress. Despite these results, the effects and mechanisms of periodic mechanical stress are still not fully understood, so in this study we extended our study using phosphoproteomic techniques...
2017: Cellular Physiology and Biochemistry
Taewook Kang, Pia Jensen, Honggang Huang, Gitte Lund Christensen, Nils Billestrup, Martin R Larsen
Normal pancreatic islet β-cells (PBCs) abundantly secrete insulin in response to elevated blood glucose levels, in order to maintain an adequate control of energy balance and glucose homeostasis. However, the molecular mechanisms underlying the insulin secretion are unclear. Improving our understanding of glucose-stimulated insulin secretion (GSIS) mechanisms under normal conditions is a prerequisite for developing better interventions against diabetes. Here, we aimed at identifying novel signaling pathways involved in the initial release of insulin from PBCs after glucose stimulation using quantitative strategies for the assessment of phosphorylated proteins and sialylated N-linked (SA) glycoproteins...
November 7, 2017: Molecular & Cellular Proteomics: MCP
Ana Contreras, Lidia Morales, Ali Tebourbi, Miguel Miguéns, Nuria Del Olmo, Carmen Pérez-García
Alcohol consumption during adolescence is deleterious to the developing brain and leads to persistent deficits in adulthood. Several results provide strong evidence for ethanol-associated alterations in glutamatergic signaling and impaired synaptic plasticity in the hippocampus. Protein phosphorylation is a well-known and well-documented mechanism in memory processes, but information on phosphoprotein alterations in hippocampus after ethanol exposure is limited. This study focuses on age-related changes in the hippocampal phosphoproteome after acute alcohol administration...
December 18, 2017: Chemical Research in Toxicology
Joana Machado Oliveira, Cristóvão B da Cruz E Silva, Thorsten Müller, Tânia Soares Martins, Marta Cova, Odete A B da Cruz E Silva, Ana Gabriela Henriques
Neuroproteomics is an evolving field of postgenomic medicine, highlighting the convergence of psychiatry/neurology and proteomics, yet compared with neurogenetics, it has received little attention. This study in rat primary neuronal cultures provides an example of a neuroproteomic approach relevant to the study of psychiatric disease pathophysiology, focusing on Alzheimer's disease. In this context, okadaic acid (OA) is routinely used in experimental designs to investigate phosphorylation-mediated events. It is a potent protein phosphatase (PP) inhibitor, particularly of PP1 and PP2A...
September 2017: Omics: a Journal of Integrative Biology
Hongbo Guo, Ruth Isserlin, Andrew Emili, Jatin G Burniston
Endurance exercise improves cardiac performance and affords protection against cardiovascular diseases but the signalling events that mediate these benefits are largely unexplored. Phosphorylation is a widely studied post-translational modification involved in intracellular signalling, and to discover novel phosphorylation events associated with exercise we have profiled the cardiac phosphoproteome response to a standardised exercise test to peak oxygen uptake (VO2peak). Male Wistar rats (346±18g) were assigned to 3 independent groups (n=6, in each) that were familiarised with running on a motorised treadmill within a metabolic chamber...
October 2017: Journal of Molecular and Cellular Cardiology
Hirohisa Nagahori, Kenji Nakamura, Kayo Sumida, Shingo Ito, Sumio Ohtsuki
Transcriptomic, proteomic, phosphoproteomic, and metabolomic analyses were combined to determine the role of pregnane X receptor (PXR) in nongenotoxic signaling and energy homeostasis in liver after rats were repeatedly orally dosed with the PXR agonist pregnenolone carbonitrile (PCN) for 7 days. Analyses of mRNAs and proteins in the supernatant, membrane, and cytosolic fractions of enlarged liver homogenates showed diverse expression profiles. Gene set enrichment analysis showed that the synchronous increase in mRNAs and proteins involved in chemical carcinogenesis and the response to drug was possibly mediated by the PXR pathway and proteasome core complex assembly was possibly mediated by the Nrf2 pathway...
September 6, 2017: Journal of Proteome Research
Yuanyuan Zhang, Yajie Zhang, Yonghao Yu
Insulin resistance is a hallmark of type 2 diabetes. Although multiple genetic and physiological factors interact to cause insulin resistance, deregulated signaling by phosphorylation is a common underlying mechanism. In particular, the specific phosphorylation-dependent regulatory mechanisms and signaling outputs of insulin are poorly understood in hepatocytes, which represents one of the most important insulin-responsive cell types. Using primary rat hepatocytes as a model system, we performed reductive dimethylation (ReDi)-based quantitative mass spectrometric analysis and characterized the phosphoproteome that is regulated by insulin as well as its key downstream kinases including Akt, mTORC1, and S6K...
August 4, 2017: Journal of Proteome Research
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
Qian Wang, Xiao-Mei He, Xi Chen, Gang-Tian Zhu, Ren-Qi Wang, Yu-Qi Feng
Phosphorylation is a crucial post-translational modification, which plays pivotal roles in various biological processes. Analysis of phosphopeptides by mass spectrometry (MS) is intractable on account of their low stoichiometry and the ion suppression from non-phosphopeptides. Thus, enrichment of phosphopeptides before MS analysis is indispensable. In this work, we employed pyridoxal 5'-phosphate (PLP), as an immobilized metal affinity chromatography (IMAC) ligand for the enrichment of phosphopeptides. PLP was grafted onto several substrates such as silica (SiO2), oxidized carbon nanotube (OCNT) and silica coated magnetic nanoparticles (Fe3O4@SiO2)...
May 26, 2017: Journal of Chromatography. A
Harm Post, Renske Penning, Martin A Fitzpatrick, Luc B Garrigues, W Wu, Harold D MacGillavry, Casper C Hoogenraad, Albert J R Heck, A F Maarten Altelaar
Because of the low stoichiometry of protein phosphorylation, targeted enrichment prior to LC-MS/MS analysis is still essential. The trend in phosphoproteome analysis is shifting toward an increasing number of biological replicates per experiment, ideally starting from very low sample amounts, placing new demands on enrichment protocols to make them less labor-intensive, more sensitive, and less prone to variability. Here we assessed an automated enrichment protocol using Fe(III)-IMAC cartridges on an AssayMAP Bravo platform to meet these demands...
February 3, 2017: Journal of Proteome Research
Jaime Santo-Domingo, Isabelle Chareyron, Loïc Dayon, Antonio Núñez Galindo, Ornella Cominetti, María Pilar Giner Giménez, Umberto De Marchi, Carles Canto, Martin Kussmann, Andreas Wiederkehr
Mitochondria play a central role in pancreatic β-cell nutrient sensing by coupling their metabolism to plasma membrane excitability and insulin granule exocytosis. Whether non-nutrient secretagogues stimulate mitochondria as part of the molecular mechanism to promote insulin secretion is not known. Here, we show that PKC signaling, which is employed by many non-nutrient secretagogues, augments mitochondrial respiration in INS-1E (rat insulinoma cell line clone 1E) and human pancreatic β cells. The phorbol ester, phorbol 12-myristate 13-acetate, accelerates mitochondrial respiration at both resting and stimulatory glucose concentrations...
March 2017: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
Shaoling Jiang, Hanchang He, Lishan Tan, Liangliang Wang, Zhengxiu Su, Yufeng Liu, Hongguo Zhu, Menghuan Zhang, Fan Fan Hou, Aiqing Li
Salt plays an essential role in the progression of chronic kidney disease and hypertension. However, the mechanisms underlying pathogenesis of salt-induced kidney damage remain largely unknown. Here, Sprague-Dawley rats, that underwent 5/6 nephrectomy (5/6Nx, a model of advanced kidney damage) or sham operation, were treated for 2 weeks with a normal or high-salt diet. We employed aTiO2 enrichment, iTRAQ labeling and liquid-chromatography tandem mass spectrometry strategy for proteomic and phosphoproteomic profiling of the renal cortex...
October 24, 2016: Scientific Reports
Matthew T Rich, Thomas B Abbott, Lisa Chung, Erol E Gulcicek, Kathryn L Stone, Christopher M Colangelo, TuKiet T Lam, Angus C Nairn, Jane R Taylor, Mary M Torregrossa
UNLABELLED: Successful addiction treatment depends on maintaining long-term abstinence, making relapse prevention an essential therapeutic goal. However, exposure to environmental cues associated with drug use often thwarts abstinence efforts by triggering drug using memories that drive craving and relapse. We sought to develop a dual approach for weakening cocaine memories through phosphoproteomic identification of targets regulated in opposite directions by memory extinction compared with reconsolidation in male Sprague-Dawley rats that had been trained to self-administer cocaine paired with an audiovisual cue...
July 20, 2016: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
Mahdokht Kohansal-Nodehi, John Je Chua, Henning Urlaub, Reinhard Jahn, Dominika Czernik
Neurotransmitter release is mediated by the fast, calcium-triggered fusion of synaptic vesicles with the presynaptic plasma membrane, followed by endocytosis and recycling of the membrane of synaptic vesicles. While many of the proteins governing these processes are known, their regulation is only beginning to be understood. Here we have applied quantitative phosphoproteomics to identify changes in phosphorylation status of presynaptic proteins in resting and stimulated nerve terminals isolated from the brains of Wistar rats...
April 26, 2016: ELife
Shuxin Wang, Zheyi Li, Hongyan Shen, Zhong Zhang, Yuxin Yin, Qingsong Wang, Xuyang Zhao, Jianguo Ji
Protein phosphorylation is central to the understanding of multiple cellular signaling pathways responsible for regulating the self-renewal and differentiation of neural stem cells (NSCs). Here we performed a large-scale phosphoproteomic analysis of rat fetal NSCs using strong cation exchange chromatography prefractionation and citric acid-assisted two-step enrichment with TiO2 strategy followed by nanoLC-MS/MS analysis. Totally we identified 32,546 phosphosites on 5,091 phosphoproteins, among which 23,945 were class I phosphosites, and quantified 16,000 sites during NSC differentiation...
August 2016: Stem Cells
Liyuan Zhang, Zhen Liang, Lihua Zhang, Yukui Zhang, Shujuan Shao
Despite recent advances in phosphoproteome research, detection and characterization of multi-phosphopeptides have remained a challenge. Here we present a novel IMAC strategy for effective extracting multi-phosphopeptides from complex samples, through Ga(3+) chelation to the adenosine tri-phosphate (ATP)-functionalized magnetic nanoparticles (Ga(3+)-ATP-MNPs). The high specificity of Ga(3+)-ATP-MNPs was demonstrated by efficient enriching multi-phosphopeptides from the digest mixture of β-casein and BSA with molar ratio as low as 1:5000...
November 5, 2015: Analytica Chimica Acta
Jiaming Li, Qingrun Li, Jiashu Tang, Fangying Xia, Jiarui Wu, Rong Zeng
As central tissue of glucose homeostasis, islet has been an important focus of diabetes research. Phosphorylation plays pivotal roles in islet function, especially in islet glucose-stimulated insulin secretion. A systematic view on how phosphorylation networks were coordinately regulated in this process remains lacking, partially due to the limited amount of islets from an individual for a phosphoproteomic analysis. Here we optimized the in-tip and best-ratio phosphopeptide enrichment strategy and a SILAC-based workflow for processing rat islet samples...
November 6, 2015: Journal of Proteome Research
Rima Chaudhuri, Arash Sadrieh, Nolan J Hoffman, Benjamin L Parker, Sean J Humphrey, Jacqueline Stöckli, Adam P Hill, David E James, Jean Yee Hwa Yang
BACKGROUND: Most biological processes are influenced by protein post-translational modifications (PTMs). Identifying novel PTM sites in different organisms, including humans and model organisms, has expedited our understanding of key signal transduction mechanisms. However, with increasing availability of deep, quantitative datasets in diverse species, there is a growing need for tools to facilitate cross-species comparison of PTM data. This is particularly important because functionally important modification sites are more likely to be evolutionarily conserved; yet cross-species comparison of PTMs is difficult since they often lie in structurally disordered protein domains...
2015: BMC Genomics
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