Proteolytic cleavage of phospholipase C–γ1 during apoptosis in Molt–4 cells

• DOI: 10.1096/fasebj.14.9.1083
• Publication Date: 2018-01-18T12:59:22Z
• Authors: Sun Sik Bae; David K Perry; Yong Seok Oh; Jang Hyun Choi; Sehamuddin H. Gal...; Tariq Ghayur; Sung Ho Ryu; Yusuf A. Hannun; Pann-Ghill Suh
• Keywords: 0301 basic medicine; 570; proteolysis; PROTEIN; BCL-2; ELEGANS; Apoptosis; Ceramides; CYTOCHROME-C; ACTIVATION; Mice; 03 medical and health sciences; Animals; Amino Acid Sequence; SH3 DOMAIN; Phosphorylation; PHOSPHORYLATION; Phosphotyrosine; PLC-gamma 1; Etoposide; Caspase 7; Caspase 3; Phospholipase C gamma; tyrosine phosphorylation; DEATH; FOCAL ADHESION; Caspase Inhibitors; ErbB Receptors; Isoenzymes; Molecular Weight; Amino Acid Substitution; Proto-Oncogene Proteins c-bcl-2; Caspases; Mutation; CASPASE FAMILY; Signal Transduction

Apoptosis is a cell suicide mechanism that requires the activation of cellular death proteases for its induction. We examined whether the progress of apoptosis involves cleavage of phospholipase C-gamma1 (PLC-gamma1), which plays a pivotal role in mitogenic signaling pathway. Pretreatment of T leukemic Molt-4 cells with PLC inhibitors such as U-73122 or ET-18-OCH(3) potentiated etoposide-induced apoptosis in these cells. PLC-gamma1 was fragmented when Molt-4 cells were treated with several apoptotic stimuli such as etoposide, ceramides, and tumor necrosis factor alpha. Cleavage of PLC-gamma1 was blocked by overexpression of Bcl-2 and by specific inhibitors of caspases such as Z-DEVD-CH(2)F and YVAD-cmk. Purified caspase-3 and caspase-7, group II caspases, cleaved PLC-gamma1 in vitro and generated a cleavage product of the same size as that observed in vivo, suggesting that PLC-gamma1 is cleaved by group II caspases in vivo. From point mutagenesis studies, Ala-Glu-Pro-Asp(770) was identified to be a cleavage site within PLC-gamma1. Epidermal growth factor receptor (EGFR) -induced tyrosine phosphorylation of PLC-gamma1 resulted in resistance to cleavage by caspase-3 in vitro. Furthermore, cleaved PLC-gamma1 could not be tyrosine-phosphorylated by EGFR in vitro. In addition, tyrosine-phosphorylated PLC-gamma1 was not significantly cleaved during etoposide-induced apoptosis in Molt-4 cells. This suggests that the growth factor-induced tyrosine phosphorylation may suppress apoptosis-induced fragmentation of PLC-gamma1. We provide evidence for the biochemical relationship between PLC-gamma1-mediated signal pathway and apoptotic signal pathway, indicating that the defect of PLC-gamma1-mediated signaling pathway can facilitate an apoptotic progression.