The proapoptotic activity of C-terminal domain of apoptosis-inducing factor (AIF) is separated from its N-terminal

• YONG ZHANG ^[1] ; TAO HAN ^[1] ; QING ZHU ^[2] ; WEI ZHANG ^[1] ; WEI BAO ^[1] ; HAI-JING FU ^[1] ; JIE YANG ^[3] ; XIAO-JUN HUANG ^[4] ; JUN-XIA WEI ^[3] ; YAN-LING MENG ^[1] ; JING ZHAO ^[3] ; YUN-XIN CAO ^[1] ; LIN-TAO JIA ^[3] ; AN-GANG YANG ^[1]
  1. [1] Fourth Military Medical University

     Fourth Military Medical University

     China

     
  2. [2] First Affiliated Hospital of Xi'an Jiaotong Universit Department of Medical Oncology
  3. [3] First Affiliated Hospital of Xi'an Jiaotong Universit Biochemistry and Molecular Biology
  4. [4] First Affiliated Hospital of Xi'an Jiaotong Universit Preclinical Medical Teaching Lab Center

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• Localización: Biological Research, ISSN-e 0717-6287, ISSN 0716-9760, Vol. 42, Nº. 2, 2009, págs. 249-260
• Idioma: inglés
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• Resumen

  • Apoptosis-inducing factor (AIF) is a mitochondrial flavoprotein that mediates both NADH-oxidizing and caspase-independent apoptosis. Further, the proapoptotic activity of AIF is located in the C-terminus of AIF, although the precise minimum sequence responsible for apoptosis induction remains to be investigated. In the present study, we generated two truncated AIFs, AIFΔ1-480-FLAG, which is a FLAG-tagged C-terminal peptide comprising amino acids from 481 to 613, and AIF360-480 containing amino acids from 360 to 480 of AIF. We used confocal microscopy to demonstrate that both the truncated proteins are expressed and located in the cytoplasm of transfected cells. AIFΔ1-480 but not AIF360-480 induces apoptosis in transfected cells. We also found that the expression of AIFΔ1-480 could initiate the release of cytochrome c from the mitochondria. The suppression of caspase-9 via siRNA blocked the proapoptotic activity of AIFΔ1-480. Therefore, AIFΔ 1-480 is sufficient for inducing caspase-9-dependent apoptotic signaling, probably by promoting the release of cytochrome c. At last, we generated a chimeric immuno-AIFΔ 1-480 protein, which comprised an HER2 antibody, a Pseudomonas exotoxin A translocation domain and AIFΔ 1-480. Human Jurkat cells transfected with the immuno-AIFΔl-480 gene could express and secrete the chimeric protein, which selectively recognize and kill HER2-overexpressing tumor cells. Our study demonstrates the feasibility of the immuno-AIFΔl-480 gene as a novel approach to treating HER2-overexpressing cancers.