scholarly journals Proteomic Analysis of Tears and Conjunctival Cells Collected with Schirmer Strips Using timsTOF Pro: Preanalytical Considerations

Metabolites ◽  
2021 ◽  
Vol 12 (1) ◽  
pp. 2
Author(s):  
Murat Akkurt Arslan ◽  
Ioannis Kolman ◽  
Cédric Pionneau ◽  
Solenne Chardonnet ◽  
Romain Magny ◽  
...  
Keyword(s):  
Ion Mobility ◽  
Biological Processes ◽  
Limited Sample ◽  
Biological Regulation ◽  
High Data ◽  
Cellular Processes ◽  
Proteomic Approach ◽  
Acquisition Speed ◽  
Conjunctival Cells ◽  
Proteome Profile

This study aimed to investigate the human proteome profile of samples collected from whole (W) Schirmer strips (ScS) and their two parts—the bulb (B) and the rest of the strip (R)—with a comprehensive proteomic approach using a trapped ion mobility mass spectrometer, the timsTOF Pro. Eight ScS were collected from two healthy subjects at four different visits to be separated into three batches, i.e., 4W, 4B, and 4R. In total, 1582 proteins were identified in the W, B, and R batches. Among all identified proteins, binding proteins (43.4%) and those with catalytic activity (42.2%) constituted more than 80% of the molecular functions. The most represented biological processes were cellular processes (31.2%), metabolic processes (20.8%), and biological regulation (13.1%). Enzymes were the most represented protein class (41%), consisting mainly of hydrolases (47.5%), oxidoreductases (22.1%), and transferases (16.7%). The bulb (B), which is in contact with the conjunctiva, might collect both tear and cell proteins and therefore promote the identification of more proteins. Processing B and R separately before mass spectrometry (MS) analysis, combined with the high data acquisition speed and the addition of ion-mobility-based separation in the timsTOF Pro, can bring a new dimension to biomarker investigations of a limited sample such as tear fluid.

scholarly journals Biophysical basis of cellular multi-specificity encoded in a model molecular switch

2020 ◽  
Author(s):  
Tina Perica ◽  
Christopher J. P. Mathy ◽  
Jiewei Xu ◽  
Gwendolyn M. Jang ◽  
Yang Zhang ◽  
...  
Keyword(s):  
Kinetic Parameters ◽  
Molecular Mechanisms ◽  
Point Mutations ◽  
Small Gtpase ◽  
Differential Sensitivity ◽  
Physical Interaction ◽  
Biological Processes ◽  
Biological Regulation ◽  
Cellular Processes ◽  
Disease Mutations

Molecular switches are central to signal transduction in protein interaction networks. One switch protein can independently regulate distinct cellular processes, but the molecular mechanisms enabling this functional multi-specificity remain unclear. Here we integrate system-scale cellular and biophysical measurements to study how a paradigm switch, the small GTPase Ran/Gsp1, achieves its functional multi-specificity. We make 56 targeted point mutations to individual interactions of Ran/Gsp1 and show through quantitative, systematic genetic and physical interaction mapping that Ran/Gsp1 interface perturbations have widespread cellular consequences that cluster by biological processes but, unexpectedly, not by the targeted interactions. Instead, the cellular consequences of the interface mutations group by their biophysical effects on kinetic parameters of the GTPase switch cycle, and cycle kinetics are allosterically tuned by distal interface mutations. We propose that the functional multi-specificity of Ran/Gsp1 is encoded by a differential sensitivity of biological processes to different kinetic parameters of the Gsp1 switch cycle, and that Gsp1 partners binding to the sites of distal mutations act as allosteric regulators of the switch. Similar mechanisms may underlie biological regulation by other GTPases and biological switches. Finally, our integrative platform to determine the quantitative consequences of cellular perturbations may help explain the effects of disease mutations targeting central switches.

scholarly journals Bayesian graphical models for modern biological applications

2021 ◽  
Author(s):  
Yang Ni ◽  
Veerabhadran Baladandayuthapani ◽  
Marina Vannucci ◽  
Francesco C. Stingo
Keyword(s):  
Graphical Models ◽  
Cancer Genomics ◽  
Graph Structure ◽  
Biological Processes ◽  
Limited Sample ◽  
Large Networks ◽  
Bayesian Approaches ◽  
Complex Sampling ◽  
Complex Dependence ◽  
Bayesian Graphical Models

AbstractGraphical models are powerful tools that are regularly used to investigate complex dependence structures in high-throughput biomedical datasets. They allow for holistic, systems-level view of the various biological processes, for intuitive and rigorous understanding and interpretations. In the context of large networks, Bayesian approaches are particularly suitable because it encourages sparsity of the graphs, incorporate prior information, and most importantly account for uncertainty in the graph structure. These features are particularly important in applications with limited sample size, including genomics and imaging studies. In this paper, we review several recently developed techniques for the analysis of large networks under non-standard settings, including but not limited to, multiple graphs for data observed from multiple related subgroups, graphical regression approaches used for the analysis of networks that change with covariates, and other complex sampling and structural settings. We also illustrate the practical utility of some of these methods using examples in cancer genomics and neuroimaging.

scholarly journals Intrinsically Disordered Proteins as Regulators of Transient Biological Processes and as Untapped Drug Targets

Molecules ◽  
2021 ◽  
Vol 26 (8) ◽  
pp. 2118
Author(s):  
Yusuke Hosoya ◽  
Junko Ohkanda
Keyword(s):  
Drug Targets ◽  
Intrinsically Disordered Proteins ◽  
Dynamic Control ◽  
Disordered Proteins ◽  
Biological Processes ◽  
Phase Separations ◽  
Cellular Processes ◽  
Intrinsically Disordered ◽  
New Drug ◽  
Liquid Phase Separations

Intrinsically disordered proteins (IDPs) are critical players in the dynamic control of diverse cellular processes, and provide potential new drug targets because their dysregulation is closely related to many diseases. This review focuses on several medicinal studies that have identified low-molecular-weight inhibitors of IDPs. In addition, clinically relevant liquid–liquid phase separations—which critically involve both intermolecular interactions between IDPs and their posttranslational modification—are analyzed to understand the potential of IDPs as new drug targets.

scholarly journals Proteomic analysis of anti-angiogenic effects by conbercept in the mice with oxygen induced retinopathy

2020 ◽  
Vol 13 (12) ◽  
pp. 1844-1853
Author(s):  
Ji Jin ◽  
Gao-Qin Liu ◽  
Pei-Rong Lu
Keyword(s):  
Risk Factors ◽  
Molecular Mechanisms ◽  
Hypoxia Inducible Factor ◽  
Fibrin Clot ◽  
Biological Regulation ◽  
Cellular Processes ◽  
Novel Proteins ◽  
Oxygen Induced Retinopathy ◽  
Single Organism ◽  
Group 2

AIM: To analyze the retinal proteomes with and without conbercept treatments in mice with oxygen-induced retinopathy (OIR) and identify proteins involved in the molecular mechanisms mediated by conbercept. METHODS: OIR was induced in fifty-six C57BL/6J mouse pups and randomly divided into four groups. Group 1: Normal17 (n=7), mice without OIR and treated with normal air. Group 2: OIR12/EXP1 (n=14), mice received 75% oxygen from postnatal day (P) 7 to 12. Group 3: OIR17/Control (n=14), mice received 75% oxygen from P7 to P12 and then normal air to P17. Group 4: Lang17/EXP2 (n=21), mice received 75% oxygen from P7 to P12 with intravitreal injection of 1 µL conbercept at the concentration of 10 mg/mL at P12, and then normal air from P12 to P17. Liquid Chromatography-Mass Spectrometry (LC-MS)/MS data were reviewed to find proteins that were up-regulated after the conbercept treatment. Gene ontology (GO) analysis was performed of conbercept-mediated changes in proteins involved in single-organism processes, biological regulation, cellular processes, immune responses, metabolic processes, locomotion and multiple-organism processes. RESULTS: Conbercept induced a reversal of hypoxia-inducible factor 1 signaling pathway as revealed by the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and also induced down-regulation of proteins involved in blood coagulation and fibrin clot formation as demonstrated by the Database for Annotation, Visualization and Integrated Discovery (DAVID) and the stimulation of interferon genes studies. These appear to be risk factors of retinal fibrosis. Additional conbercept-specific fibrosis risk factors were also identified and may serve as therapeutic targets for fibrosis. CONCLUSION: Our studies reveal that many novel proteins are differentially regulated by conbercept. The new insights may warrant a valuable resource for conbercept treatment.

Comparative whey proteome analysis of small-tailed Han and DairyMeade ovine milk

2021 ◽  
pp. 1-5
Author(s):  
Urhan Bai ◽  
Xiaohu Su ◽  
Zhong Zheng ◽  
Liguo Zhang ◽  
Ying Ma ◽  
...  
Keyword(s):  
Mammary Gland Development ◽  
Proteome Analysis ◽  
Dairy Sheep ◽  
Milk Traits ◽  
Beneficial Effects ◽  
Proteomic Approach ◽  
Tandem Mass Tag ◽  
Proteome Profile ◽  
Related Proteins ◽  
Gland Development

Abstract We characterized the proteome profile of mid-lactation small-tailed Han (STH) and DairyMeade (DM) ovine milk in order to explore physiological variation and differences in milk traits between the two breeds. Methodology combined a tandem mass tag (TMT) proteomic approach with LC-MS/MS technology. A total of 656 proteins were identified in STH and DM ovine milk, of which 17and 29 proteins were significantly upregulated (P < 0.05) in STH and DM, respectively. Immune-related proteins and disease-related proteins were highly expressed in STH milk, whereas S100A2 and AEBP1 were highly expressed in DM milk, which had beneficial effects on mammary gland development and milk yield. Our results provide a theoretical basis for future breeding of dairy sheep.

scholarly journals circStrn3 is involved in bone cancer pain regulation in a rat model

2020 ◽  
Vol 52 (5) ◽  
pp. 495-505
Author(s):  
Yiwen Zhang ◽  
Xiaoxia Zhang ◽  
Zumin Xing ◽  
Shuyi Tang ◽  
Hanwen Chen ◽  
...  
Keyword(s):  
Cancer Pain ◽  
Rat Model ◽  
Molecular Mechanisms ◽  
Bone Cancer ◽  
Test Body ◽  
Differentially Expressed ◽  
Bone Cancer Pain ◽  
Biological Regulation ◽  
Cellular Processes ◽  
Simplex Infection

Abstract Bone cancer pain (BCP) is a common chronic pain that is caused by a primary or metastatic bone tumor. More detailed molecular mechanisms of BCP are warranted. In this study, we established a BCP rat model. The von Frey hair test, body weight, and hematoxylin and eosin staining were employed. We screened differentially expressed circRNAs (DECs) between the BCP group and sham group. The results revealed that 850 DECs were significantly up-regulated and 644 DECs were significantly down-regulated in the BCP group. Furthermore, we identified 1177 differentially expressed genes (DEGs) significantly up-regulated and 565 DEGs significantly down-regulated in the BCP group. Gene Ontology annotation of all 1742 DEGs revealed that biological regulation of metabolic processes, cellular processes, and binding were the top enriched terms. For Kyoto Encyclopedia of Genes and Genomes analysis, phagosome, HTLV-I infection, proteoglycans in cancer, and herpes simplex infection were significantly enriched in this study. In addition, we identified four selected circRNAs, chr6:72418120|72430205, chr20:7561057|7573740, chr18:69943105|69944476, and chr5:167516581|167558250, by quantitative real time PCR. chr6:72418120|72430205 (circStrn3) was selected for further study based on expression level and the circRNA–miRNA–mRNA network table. Western blot analysis suggested that knockdown of circStrn3 could effectively induce Walker 256 cell apoptosis. In summary, our study provided a more in-depth understanding of the molecular mechanisms of BCP.

Nanomechanochemical Delivery of Nanoparticles for Nanomechanics Inside Living Cells

2010 ◽  
Author(s):  
Kyungsuk Yum ◽  
Sungsoo Na ◽  
Yang Xiang ◽  
Ning Wang ◽  
Min-Feng Yu
Keyword(s):  
Single Molecule ◽  
Living Cells ◽  
Endocytic Pathway ◽  
Great Promise ◽  
Biological Processes ◽  
Temporal Precision ◽  
Single Molecule Level ◽  
Cellular Processes ◽  
Cellular Environment ◽  
Biological Studies

Studying biological processes and mechanics in living cells is challenging but highly rewarding. Recent advances in experimental techniques have provided numerous ways to investigate cellular processes and mechanics of living cells. However, most of existing techniques for biomechanics are limited to experiments outside or on the membrane of cells, due to the difficulties in physically accessing the interior of living cells. On the other hand, nanomaterials, such as fluorescent quantum dots (QDs) and magnetic nanoparticles, have shown great promise to overcome such limitations due to their small sizes and excellent functionalities, including bright and stable fluorescence and remote manipulability. However, except a few systems, the use of nanoparticles has been limited to the study of biological studies on cell membranes or related to endocytosis, because of the difficulty of delivering dispersed and single nanoparticles into living cells. Various strategies have been explored, but delivered nanoparticles are often trapped in the endocytic pathway or form aggregates in the cytoplasm, limiting their further use. Here we show a nanoscale direct delivery method, named nanomechanochemical delivery, where we manipulate a nanotube-based nanoneedle, carrying “cargo” (QDs in this study), to mechanically penetrate the cell membrane, access specific areas inside cells, and release the cargo [1]. We selectively delivered well-dispersed QDs into either the cytoplasm or the nucleus of living cells. We quantified the dynamics of the delivered QDs by single-molecule tracking and demonstrated the applicability of the QDs as a nanoscale probe for studying nanomechanics inside living cells (by using the biomicrorhology method), revealing the biomechanical heterogeneity of the cellular environment. This method may allow new strategies for studying biological processes and mechanics in living cells with spatial and temporal precision, potentially at the single-molecule level.

scholarly journals Proteomic Analysis of Mucopolysaccharidosis IIIB Mouse Brain

Biomolecules ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 355 ◽  
Author(s):  
Valeria De Pasquale ◽  
Michele Costanzo ◽  
Rosa Siciliano ◽  
Maria Mazzeo ◽  
Valeria Pistorio ◽  
...  
Keyword(s):  
Mouse Brain ◽  
Clinical Manifestations ◽  
Label Free ◽  
Wild Type ◽  
Proteomic Approach ◽  
Cytoskeletal Regulation ◽  
Proteome Profile ◽  
Subsequent Storage ◽  
And Control ◽  
Differentially Abundant Proteins

Mucopolysaccharidosis IIIB (MPS IIIB) is an inherited metabolic disease due to deficiency of α-N-Acetylglucosaminidase (NAGLU) enzyme with subsequent storage of undegraded heparan sulfate (HS). The main clinical manifestations of the disease are profound intellectual disability and neurodegeneration. A label-free quantitative proteomic approach was applied to compare the proteome profile of brains from MPS IIIB and control mice to identify altered neuropathological pathways of MPS IIIB. Proteins were identified through a bottom up analysis and 130 were significantly under-represented and 74 over-represented in MPS IIIB mouse brains compared to wild type (WT). Multiple bioinformatic analyses allowed to identify three major clusters of the differentially abundant proteins: proteins involved in cytoskeletal regulation, synaptic vesicle trafficking, and energy metabolism. The proteome profile of NAGLU−/− mouse brain could pave the way for further studies aimed at identifying novel therapeutic targets for the MPS IIIB. Data are available via ProteomeXchange with the identifier PXD017363.

scholarly journals Small RNA Expression Profiling by High-Throughput Sequencing: Implications of Enzymatic Manipulation

2012 ◽  
Vol 2012 ◽  
pp. 1-15 ◽  
Author(s):  
Fanglei Zhuang ◽  
Ryan T. Fuchs ◽  
G. Brett Robb
Keyword(s):  
High Throughput ◽  
Expression Profiling ◽  
Messenger Rna ◽  
High Throughput Sequencing ◽  
Biological Processes ◽  
Cellular Processes ◽  
Disease States ◽  
Powerful Approach ◽  
Sequencing Library ◽  
Rna Expression Profiling

Eukaryotic regulatory small RNAs (sRNAs) play significant roles in many fundamental cellular processes. As such, they have emerged as useful biomarkers for diseases and cell differentiation states. sRNA-based biomarkers outperform traditional messenger RNA-based biomarkers by testing fewer targets with greater accuracy and providing earlier detection for disease states. Therefore, expression profiling of sRNAs is fundamentally important to further advance the understanding of biological processes, as well as diagnosis and treatment of diseases. High-throughput sequencing (HTS) is a powerful approach for both sRNA discovery and expression profiling. Here, we discuss the general considerations for sRNA-based HTS profiling methods from RNA preparation to sequencing library construction, with a focus on the causes of systematic error. By examining the enzymatic manipulation steps of sRNA expression profiling, this paper aims to demystify current HTS-based sRNA profiling approaches and to aid researchers in the informed design and interpretation of profiling experiments.

scholarly journals Systematic identification of signal integration by protein kinase A

2015 ◽  
Vol 112 (14) ◽  
pp. 4501-4506 ◽  
Author(s):  
Marie Filteau ◽  
Guillaume Diss ◽  
Francisco Torres-Quiroz ◽  
Alexandre K. Dubé ◽  
Andrea Schraffl ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Protein Interactions ◽  
Carbon Sources ◽  
Biological Processes ◽  
Protein Protein Interactions ◽  
Cellular Processes ◽  
Protein Complex Formation ◽  
Growth And Aging ◽  
Kinase A

Cellular processes and homeostasis control in eukaryotic cells is achieved by the action of regulatory proteins such as protein kinase A (PKA). Although the outbound signals from PKA directed to processes such as metabolism, growth, and aging have been well charted, what regulates this conserved regulator remains to be systematically identified to understand how it coordinates biological processes. Using a yeast PKA reporter assay, we identified genes that influence PKA activity by measuring protein–protein interactions between the regulatory and the two catalytic subunits of the PKA complex in 3,726 yeast genetic-deletion backgrounds grown on two carbon sources. Overall, nearly 500 genes were found to be connected directly or indirectly to PKA regulation, including 80 core regulators, denoting a wide diversity of signals regulating PKA, within and beyond the described upstream linear pathways. PKA regulators span multiple processes, including the antagonistic autophagy and methionine biosynthesis pathways. Our results converge toward mechanisms of PKA posttranslational regulation by lysine acetylation, which is conserved between yeast and humans and that, we show, regulates protein complex formation in mammals and carbohydrate storage and aging in yeast. Taken together, these results show that the extent of PKA input matches with its output, because this kinase receives information from upstream and downstream processes, and highlight how biological processes are interconnected and coordinated by PKA.

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