Activation of T lymphoma cells expressing Syk, but not ZAP-70 tyrosine kinase, has been shown to negatively regulate cell activation and activation-induced cell death (AICD), perhaps due to differential induction of tyrosine phosphorylation modified proteins. To better understand the role of these proteins and their associated molecules/pathways, we studied a previously described model of T lymphoma cells expressing either a kinase-activated chimeric Syk or ZAP-70 genetically linked to T-cell receptor (TCR) ζ chain (Z/Syk or Z/ZAP cells, respectively). To help identify molecules and pathways linked to cell activation or AICD, a comparative semi-quantitative proteomics-based approach was utilized to analyze tyrosine-phosphorylated protein immunoprecipitates from two-minute short-term activated Z/Syk or Z/ZAP cells. Using the resulting bioinformatics data-sets, we identified several differentially immunoprecipitated proteins that could be validated biochemically. More tyrosine-phosphorylated and phosphotyrosine-associated proteins were found in Z/Syk than in Z/ZAP cells. Proteins involved in different unique functional pathways were induced in these cells and showed altered intermolecular interactions in varied pathways. Remarkably, 41% of differentially identified proteins in Z/Syk cells belonged to cell cycle or vesicle/trafficking pathways. In contrast, 21% of such proteins in Z/ZAP cells belonged to metabolism pathways. Therefore, molecular pathways involved in post-translational modifications linked to distinct cellular/physiological functions are differentially activated, which may contribute to varied activation and AICD responses of these cells. In summary, we identified proteins belonging to novel differentially activated pathways involved in TCR-mediated signaling, which may be targets for regulating activation and AICD of T lymphoma cells and for potential cancer therapy.