Figure 3: LAT encodes downstream pathway balance.

(A) Models of LAT with interacting proteins which mediate intracellular signaling pathways controlling chromatin, RNA, or protein features. The complex of LAT with interactors (“signalosome”) could control downstream pathways in a modular or coordinated fashion. In a modular model, mutation in one region of LAT and disruption of a particular interactor will disrupt one downstream pathway while leaving others active. In a coordinated model, mutation in one region of LAT which disrupts a particular interactor may disrupt other interactions or pathway activities (either directly through higher-order physical interactions or indirectly through signal cross-talk). (B) Expected results for the models proposed in (A). Modular or coordinated signaling will exhibit distinct patterns of mutant effects on pairs of pathway activities measured in the screen. (C) Scatter plot of the accessibility of two chromatin features (inferred TF activity, averaged across cells expressing a particular ORF) representing central pathways of T cell activation. FDR from permutation sampling test. ORFs supported by at least 50 cells are displayed. (D) Scatter plot of the same data as in (C), with ORFs labeled as exhibiting balanced or biased defects across AP1 and NFAT pathways. Balanced defects exhibit statistically significant defects in both AP1 and NFAT pathways. (E) Heatmap of inferred TF activity for TFs representing motif families that increase in T cell activation. ORFs (columns) are ordered by chromatin activation score. (F) Scatter plot of AP1 and NFAT TF activity in primary human CD4+ central memory T cells. Error bars represent standard deviation across replicates, p-value from KS test of single-cell values.