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Identification of DELE, a novel DAP3-binding protein which is crucial for death receptor-mediated apoptosis induction

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Abstract

Death associated protein 3 (DAP3) is known to be a highly conserved protein, and is responsible for regulating apoptosis induced by various stimuli. To understand the molecular mechanism of how DAP3 induces apoptosis, we performed yeast two-hybrid screening, and identified a novel DAP3-binding protein termed death ligand signal enhancer (DELE). In this report, we show that DELE actually binds to DAP3 in mammalian cells. We found that the cells stably expressing DELE are susceptible to apoptosis induction by the stimulation of TNF-α and TRAIL. In addition, knockdown of DELE expression rescued the HeLa cells from apoptosis induction by these stimuli. Moreover, activation of caspase-3, caspase-8 and caspase-9 induced by stimulation of TNF-α, anti-Fas or TRAIL was significantly inhibited by the knockdown of DELE expression. These results demonstrated the biological significance of DELE for apoptosis signal mediated by death receptors.

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Abbreviations

DAP3:

Death associated protein 3

DELE:

Death ligand signal enhancer

DR:

Death receptor

TNF:

Tumor necrosis factor

TRAIL:

TNF-related apoptosis inducing ligand

DD:

Death domain

DED:

Death effector domain

FADD:

Fas-associated death domain protein

CARD:

Caspase-recruitment domain

EGFP:

Enhanced green fluorescent protein

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Acknowledgment

We would like to thank Prof. Toshimitsu Uede (Department of Molecular Immunology, Institute for Genetic Medicine, Hokkaido University) whose consultations helped us in our research.

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Authors and Affiliations

  1. Department of Bioresources, Hokkaido University Research Center for Zoonosis Control, North 20, West 10, Kita-ku, Sapporo, Hokkaido, 001-0020, Japan

    Tanenobu Harada, Atsushi Iwai & Tadaaki Miyazaki

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  1. Tanenobu Harada
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  2. Atsushi Iwai
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  3. Tadaaki Miyazaki
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Corresponding author

Correspondence to Tadaaki Miyazaki.

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10495_2010_519_MOESM1_ESM.tif

Supplemental Fig. 1. Induction of apoptosis to the HeLa cells by transiently overexpressing DELE. HeLa cells were seeded onto 6-well plate, and subsequently the cells were transfected with 4 μg of control vector or expression vector of DELE after 48 h, the cells were observed using phase-contrast microscope (Eclipse TE200-U; Nikon, Tokyo, Japan). (TIFF 14937 kb)

10495_2010_519_MOESM2_ESM.tif

Supplemental Fig. 2. Rescue of the HeLa cells from the TRAIL induced apoptosis by knockdown of DELE. The HeLa cells were transfected with DELE specific siRNA or control siRNA. After 48 h, cells were stimulated with TRAIL at the series of concentrations indicated in the figure. After a 24 h post-stimulation incubation period, the cells were observed using phase-contrast microscope (Eclipse TE200-U; Nikon). (TIFF 29674 kb)

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Harada, T., Iwai, A. & Miyazaki, T. Identification of DELE, a novel DAP3-binding protein which is crucial for death receptor-mediated apoptosis induction. Apoptosis 15, 1247–1255 (2010). https://doi.org/10.1007/s10495-010-0519-3

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  • DOI: https://doi.org/10.1007/s10495-010-0519-3

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