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A Natural L-Arginine Analog, L-Canavanine-Induced Apoptosis is Suppressed by Protein Tyrosine Kinase p56lck in Human Acute Leukemia Jurkat T Cells

인체 급성백혈병 Jurkat T 세포에 있어서 L-canavanine에 의해 유도되는 세포자살기전에 미치는 단백질 티로신 키나아제 p56lck의 저해 효과

  • Park, Hae-Sun (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Jun, Do-Youn (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Woo, Hyun-Ju (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Rue, Seok-Woo (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Kim, Sang-Kook (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Kim, Kyung-Min (School of Applied Ecological Resources, College of Ecology and Environmental Sciences, Kyungpook National University) ;
  • Park, Wan (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Moon, Byung-Jo (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University) ;
  • Kim, Young-Ho (School of Life Science and Biotechnology, College of Natural Sciences, Kyungpook National University)
  • 박해선 (경북대학교 생명과학부) ;
  • 전도연 (경북대학교 생명과학부) ;
  • 우현주 (경북대학교 생명과학부) ;
  • 류석우 (경북대학교 생명과학부) ;
  • 김경민 (대구경북대학교 상주캠퍼스, 생태자원응용학부) ;
  • 김상국 (경북대학교 생명과학부) ;
  • 박완 (경북대학교 생명과학부) ;
  • 문병조 (경북대학교 생명과학부) ;
  • 김영호 (경북대학교 생명과학부)
  • Published : 2009.11.30

Abstract

To elucidate further the antitumor effects of a natural L-arginine analogue, L-canavanine, the mechanism underlying apoptogenic activity of L-canavanine and its modulation by protein tyrosine kinase $p56^{lck}$ was investigated in human Jurkat T cells. When the cells were treated with 1.25 to 2.5 mM L-canavanine for 36 h, several apoptotic events including mitochondrial membrane potential (${\Delta\Psi}m$) loss, activation of caspase-9, -3, -8, and -7, poly (ADP-ribose) polymerase (PARP) degradation, and DNA fragmentation were induced without alteration in the levels of Fas or FasL. These apoptotic changes were more significant in $p56^{lck}$-deficient Jurkat clone JCaM1.6 than in $p56^{lck}$-positive Jurkat clone E6.1. The L-canavanine-induced apoptosis observed in $p56^{lck}$-deficient JCaM1.6 cells was significantly reduced by introducing $p56^{lck}$ gene into JCaM1.6 cells by stable transfection. Treatment of JCaM1.6/lck cells with L-canavanine caused a transient 1.6-fold increase in the kinase activity of $p56^{lck}$. Both FADD-positive wild-type Jurkat T cell clone A3 and FADD-deficient Jurkat T cell clone I2.1 exhibited a similar susceptibility to the cytotoxicity of L-canavanine, excluding involvement of Fas/FasL system in triggering L-canavanine-induced apoptosis. The L-canavanine-induced apoptotic sub-$G_1$ peak and activation of caspase-3, -8, and -7 were abrogated by pan-caspase inhibitor (z-VAD-fmk), whereas L-canavanine-induced activation of caspase-9 was not affected. These results demonstrated that L-canavanine caused apoptosis of Jurkat T cells via the loss of ${\Delta\Psi}m$, and the activation of caspase-9, -3, -8, and -7, leading to PARP degradation, and that the $p56^{lck}$ kinase attenuated the ${\Delta\Psi}m$ loss and activation of caspases, and thus contributed as a negative regulator to L-canavanine-induced apoptosis.

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