Figure 4.

Composition and Dynamics of the Human PD-1 Interactome following Physiological T Cell-APC Interaction

(A) Overview of AP-MS analysis of PD-1 and PD-1-OST expressing Jurkat T cells stimulated with Raji B cells expressing or not PD-L1 and in the presence or absence of SEE. The various T cell-B cell pellets were lysed after been kept in contact for 2 and 5 min, followed by affinity purification of PD-1-OST protein complexes.

(B) Volcano plot showing proteins significantly enriched in Jurkat-PD-1^OST cells compared with Jurkat cells expressing similar levels of untagged PD-1 proteins at 2 min after stimulation with Raji-PD-L1 cells + SEE.

(C) Volcano plot showing proteins significantly enriched in Jurkat-PD-1^OST cells 2 min after stimulation with Raji-PD-L1 cells + SEE compared with Jurkat-PD-1^OST cells stimulated for 2 min with Raji cells + SEE.

(D) Volcano plot showing proteins significantly enriched in Jurkat-PD-1^OST cells 5 min after stimulation with Raji-PD-L1 cells + SEE compared with Raji-PD-L1 cells without SEE stimulation. In (B), (C), and (D), the PD-1/PDCD1, PD-L1/CD274, SHP-2/PTPN11, SHP-1/PTPN6 proteins, and Twin-Strep-tag peptide (Streptag) are shown in red, and the x and y axes show the average fold change (log₂FC) in protein abundance and the statistical significance, respectively.

(E) Intensity for selected interactors across the different replicates. Mean ± SEM is depicted (black), and individual values are shown as gray dots.

(F) Heatmap depicting PD-1 interactor intensity across time points (row-normalized to the maximal value). The iBAQ column shows the stoichiometry of interaction of each prey with the PD-1 bait at 2 min after stimulation with Raji-PD-L1 cells + SEE.