Quantitative proteomic analysis of A549 cells infected with human respiratory syncytial virus.

Molecular & Cellular Proteomics : MCP
Diane C MundayJulian A Hiscox

Abstract

Human respiratory syncytial virus (HRSV) is a major cause of pediatric lower respiratory tract disease to which there is no vaccine or efficacious chemotherapeutic strategy. Although RNA synthesis and virus assembly occur in the cytoplasm, HRSV is known to induce nuclear responses in the host cell as replication alters global gene expression. Quantitative proteomics was used to take an unbiased overview of the protein changes in transformed human alveolar basal epithelial cells infected with HRSV. Underpinning this was the use of stable isotope labeling with amino acids in cell culture coupled to LC-MS/MS, which allowed the direct and simultaneous identification and quantification of both cellular and viral proteins. To reduce sample complexity and increase data return on potential protein localization, cells were fractionated into nuclear and cytoplasmic extracts. This resulted in the identification of 1,140 cellular proteins and six viral proteins. The proteomics data were analyzed using Ingenuity Pathways Analysis to identify defined canonical pathways and functional groupings. Selected data were validated using Western blot, direct and indirect immunofluorescence confocal microscopy, and functional assays. The study served ...Continue Reading

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Methods Mentioned

BETA
electrophoresis
FCS
confocal microscopy
reverse transcription-PCR
PCR
transfection
Assay
protein folding
transgenic
protein exchange

Software Mentioned

Mascot
Ingenuity Pathways Analysis
MaxQuant
Ingenuity Pathways
IPA
Ingenuity

Related Concepts

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