scholarly journals Evaluating the Performance of New Approaches to Spot Quantification and Differential Expression in 2-Dimensional Gel Electrophoresis Studies

2010 ◽  
Vol 9 (1) ◽  
pp. 595-604 ◽  
Author(s):  
Jeffrey S. Morris ◽  
Brittan N. Clark ◽  
Wei Wei ◽  
Howard B. Gutstein
Keyword(s):  
Differential Expression ◽  
Gel Electrophoresis ◽  
New Approaches ◽  
Spot Quantification

scholarly journals Differential mitochondrial proteomic analysis of A549 cells infected with avian influenza virus subtypes H5 and H9

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Yuting Yang ◽  
Yun Zhang ◽  
Changcheng Yang ◽  
Fang Fang ◽  
Ying Wang ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Avian Influenza ◽  
Differential Expression ◽  
Gel Electrophoresis ◽  
A549 Cells ◽  
Infected Group ◽  
Differentially Expressed ◽  
Mitochondrial Proteins ◽  
Differentially Expressed Proteins ◽  
Pathogenic Avian Influenza

Abstract Background Both the highly pathogenic avian influenza (HPAI) H5N1 and low pathogenic avian influenza (LPAI) H9N2 viruses have been reported to cross species barriers to infect humans. H5N1 viruses can cause severe damage and are associated with a high mortality rate, but H9N2 viruses do not cause such outcomes. Our purpose was to use proteomics technology to study the differential expression of mitochondrial-related proteins related to H5N1 and H9N2 virus infections. Methods According to the determined viral infection titer, A549 cells were infected with 1 multiplicity of infection virus, and the mitochondria were extracted after 24 h of incubation. The protein from lysed mitochondria was analyzed by the BCA method to determine the protein concentration, as well as SDS-PAGE (preliminary analysis), two-dimensional gel electrophoresis, and mass spectrometry. Differential protein spots were selected, and Western blotting was performed to verify the proteomics results. The identified proteins were subjected to GO analysis for subcellular localization, KEGG analysis for functional classification and signaling pathways assessment, and STRING analysis for functional protein association network construction. Results In the 2-D gel electrophoresis analysis, 227 protein spots were detected in the H5N1-infected group, and 169 protein spots were detected in the H9N2-infected group. Protein spots were further subjected to mass spectrometry identification and removal of redundancy, and 32 differentially expressed proteins were identified. Compared with the H9N2 group, the H5N1-infected group had 16 upregulated mitochondrial proteins and 16 downregulated proteins. The differential expression of 70-kDa heat shock protein analogs, short-chain enoyl-CoA hydratase, malate dehydrogenase, and ATP synthase was verified by Western blot, and the results were consistent with the proteomics findings. Functional analysis indicated that these differentially expressed proteins were primarily involved in apoptosis and metabolism. Conclusions Compared with their expression in the H9N2 group, the differential expression of eight mitochondrial proteins in the H5N1 group led to host T cell activation, antigen presentation, stress response, ATP synthesis and cell apoptosis reduction, leading to higher pathogenicity of H5N1 than H9N2.

GENETIC ANALYSIS OF RESISTANCE TO BENZIMIDAZOLES IN PHYSARUM: DIFFERENTIAL EXPRESSION OF β-TUBULIN GENES

Genetics ◽  
1984 ◽  
Vol 108 (1) ◽  
pp. 123-141
Author(s):  
Timothy G Burland ◽  
Tim Schedl ◽  
Keith Gull ◽  
William F Dove
Keyword(s):  
Genetic Analysis ◽  
Differential Expression ◽  
Electrophoretic Mobility ◽  
Structural Gene ◽  
Gel Electrophoresis ◽  
Vegetative Cell ◽  
Cell Types ◽  
Wild Type ◽  
Tubulin Genes ◽  
Β Tubulin

ABSTRACT Physarum displays two vegetative cell types, uninucleate myxamoebae and multinucleate plasmodia. Mutant myxamoebae of Physarum resistant to the antitubulin drug methylbenzimidazole-2-yl-carbamate (MBC) were isolated. All mutants tested were cross-resistant to other benzimidazoles but not to cycloheximide or emetine. Genetic analysis showed that mutation to MBC resistance can occur at any one of four unlinked loci, benA, benB, benC or benD. MBC resistance of benB and benD mutants was expressed in plasmodia, but benA and benC mutant plasmodia were MBC sensitive, suggesting that benA and benC encode myxamoeba-specific products. Myxamoebae carrying the recessive benD210 mutation express a β-tubulin with noval electrophoretic mobility, in addition to a β-tubulin with wild-type mobility. This and other evidence indicates that benD is a structural gene for β-tubulin, and that at least two β-tubulin genes are expressed in myxamoebae. Comparisons of the β-tubulins of wildtype and benD210 strains by gel electrophoresis revealed that, of the three (or more) β-tubulin genes expressed in Physarum, one, benD, is expressed in both myxamoebae and plasmodia, one is expressed specifically in myxamoebae and one is expressed specifically in plasmodia. However, mutation in only one gene, benD, is sufficient to confer MBC resistance on both myxamoebae and plasmodia.

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