Differential Protein Abundance in THP-1 Macrophages Infected with Wild-Type and YopJC172A Yersinia pseudotuberculosis
Description
This dataset contains mass spectrometry-based proteomic profiles of THP-1 macrophages infected with either wild-type Yersinia pseudotuberculosis or a catalytically inactive YopJC172A mutant. The study aims to identify differentially abundant host proteins in response to YopJ activity during infection. Samples were analyzed using Orbitrap Fusion mass spectrometry following SDS-PAGE and in-gel trypsin digestion. These data provide insights into YopJ-dependent modulation of host cell signaling pathways.
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Steps to reproduce
THP-1 cells were infected for 2 hours with Yersinia pseudotuberculosis wild-type or the YopJ^C172A mutant at a multiplicity of infection (MOI) of 50:1. Uninfected cells treated identically served as controls. Following infection, cells were harvested by scraping into cold PBS and centrifuged at 500 × g for 10 minutes at 4°C. Cell pellets were stored at –80°C until lysis. Cells were lysed in RIPA buffer supplemented with a Pierce protease inhibitor cocktail tablet for 30 minutes on ice. Lysates were then centrifuged at 18,000 × g for 10 minutes at 4°C to isolate the soluble protein fraction. Three independent biological replicates were used for this experiment. Proteins were separated by SDS-PAGE, and an entire gel lane was subjected to in-gel trypsin digestion, followed by C18 ZipTip purification. Mass spectrometric analysis was performed using a 250-mm ultrahigh-performance liquid chromatography system coupled to an Orbitrap Fusion mass spectrometer (Thermo Scientific).
Institutions
- University of Florida