Analysis of CD5 interactome in primary mouse CD4+ T cells

CD4+ T cells were isolated from gene-targeted mouse expressing a One-Strep-tag (OST) at the C terminus of the CD5 protein, briefly expanded in vitro and we analyzed the interactome of their OST-tagged CD5 protein. Affinity purification of the OST tagged protein was performed using Streptactin beads, from T cells either non-stimulated, stimulated 30s, 120s or 300s with anti-CD3 and anti-CD4 antibodies, or 300s with pervanadate. Each AP-MS purification is associated with a corresponding control (purification from CD4+ T cells isolated from WT mice, cultured and stimulated in the same conditions). The number of replicate biological experiments was n=3 for all conditions (time-points), and each sample was analyzed once by single-run nanoLC-MS, resulting in 30 raw files.