Fig 2: Knockdown of ZAP70 does not affect survival of T/NK Lymphoma cell lines.(A) KHYG cell line was treated with si Non Targeting Control (NT) or siZAP70 and immunoblot analysis of the protein expression of phosphorylated ZAP-70, total Zap-70 phosphorylated LAT, total LAT, phosphorylated SLP-76, total SLP-76, phosphorylated p38 MAPk, total p38MAPK were evaluated with Actin as the loading control. (B) KHYG cells which have been either treated with the siNT or si-ZAP-70 was evaluated for apoptosis by Annexin V PI staining. Obatoclax is a positive control demonstrating induction of apoptosis. (C) ZAP70 siRNA knockdown was performed in a wider range of cell lines. Protein expression in WT and ZAP70 knockdown TCL cells (JURKAT and KARPAS-299) and NKTL cells (KHYG1, HANK-1, NK-YS, SNK-1 and SNK-6) cell lines were analysed by immunoblotting. ß- ACTIN was used as a loading control. (D) ZAP70 mRNA was measured in control and ZAP70 siRNA conditions by RT-PCR in TCL cells (JURKAT and KARPAS-299) and NKTL cells (KHYG1, HANK-1, NK-YS, SNK-1 and SNK-6) (E) TCL cells (JURKAT and KARPAS-299) and NKTL cells (KHYG1, HANK-1, NK-YS, SNK-1 and SNK-6) were stained with Annexin V-APC and PI and analyzed by flow cytometry. Obatoclax 5µM treatment is a positive control demonstrating induction of apoptosis. (F) KHYG1cells which have been either treated with the siNT or si-ZAP-70 was evaluated by Cell-Titer Glo (Promega) after 48h.