Engineered FADD Induces Apoptosis Via an Artificial Death-Inducing Signaling Complex (DISC)

Overview

Journal Int J Biomed Sci

Specialty General Medicine

Date 2013 May 16

PMID 23675143

Authors

Emiko Suzuki

Tomoki Takashina

Manabu Nakayama

Affiliations

Soon will be listed here.

Abstract

An engineered Fas-associated death domain protein (FADD), 2DEDplusE-made previously by fusing the tandem DEDs of FADD to the E protein of lambda phage-greatly enhances apoptosis-inducing activity in adherent cells in vitro. To investigate the mechanism of apoptosis-inducing activity of this engineered FADD, we compared the apoptosis-inducing activity of various other engineered FADDs. The tandem DED of 2DEDplusE contributed most to the enhancement of apoptosis, and the E protein contributed moderately. The engineered factor produced artificial death-inducing signaling complex (DISC)-like signals in the cytoplasm that appear as grains under fluorescence microscopy. Membrane blebbing associated with apoptosis was observed just after formation of grain-like signals. Immunoprecipitation analysis demonstrated that 2DEDplusE-FLAG can bind p43/p41 forms of caspase 8 but E protein-FLAG cannot. Gel filtration analysis demonstrated that 2DEDplusE forms a large complex containing partially cleaved procaspase 8 (p43/p41) in the cytoplasm; the size of this complex varies greatly. In the absence of an extrinsic signal, the engineered FADD formed artificial DISC in the cytoplasm, and then its tandem DED activated procaspase 8, which in turn executed apoptosis. The engineered FADD complex closely mimicked intrinsic DISC and increased apoptosis-inducing activity.

References

Walczak H, Haas T . Biochemical analysis of the native TRAIL death-inducing signaling complex. Methods Mol Biol. 2008; 414:221-39. DOI: 10.1007/978-1-59745-339-4_16. View

Takashina T, Nakayama M . Revival of apoptotic cells that display early-stage dynamic membrane blebbing. FEBS Lett. 2007; 581(23):4479-84. DOI: 10.1016/j.febslet.2007.08.033. View

Muppidi J, Lobito A, Ramaswamy M, Yang J, Wang L, Wu H . Homotypic FADD interactions through a conserved RXDLL motif are required for death receptor-induced apoptosis. Cell Death Differ. 2006; 13(10):1641-50. DOI: 10.1038/sj.cdd.4401855. View

Chinnaiyan A, ORourke K, Tewari M, Dixit V . FADD, a novel death domain-containing protein, interacts with the death domain of Fas and initiates apoptosis. Cell. 1995; 81(4):505-12. DOI: 10.1016/0092-8674(95)90071-3. View

Carrington P, Sandu C, Wei Y, Hill J, Morisawa G, Huang T . The structure of FADD and its mode of interaction with procaspase-8. Mol Cell. 2006; 22(5):599-610. DOI: 10.1016/j.molcel.2006.04.018. View

Stupack D, Teitz T, Potter M, Mikolon D, Houghton P, Kidd V . Potentiation of neuroblastoma metastasis by loss of caspase-8. Nature. 2006; 439(7072):95-9. DOI: 10.1038/nature04323. View

Yamagata K, Daitoku H, Shimamoto Y, Matsuzaki H, Hirota K, Ishida J . Bile acids regulate gluconeogenic gene expression via small heterodimer partner-mediated repression of hepatocyte nuclear factor 4 and Foxo1. J Biol Chem. 2004; 279(22):23158-65. DOI: 10.1074/jbc.M314322200. View

Thiede B, Rudel T . Proteome analysis of apoptotic cells. Mass Spectrom Rev. 2004; 23(5):333-49. DOI: 10.1002/mas.10079. View

Hlaing T, Guo R, Dilley K, Loussia J, Morrish T, Shi M . Molecular cloning and characterization of DEFCAP-L and -S, two isoforms of a novel member of the mammalian Ced-4 family of apoptosis proteins. J Biol Chem. 2000; 276(12):9230-8. DOI: 10.1074/jbc.M009853200. View

10.

Dokland T, Murialdo H . Structural transitions during maturation of bacteriophage lambda capsids. J Mol Biol. 1993; 233(4):682-94. DOI: 10.1006/jmbi.1993.1545. View

11.

Kischkel F, Hellbardt S, Behrmann I, Germer M, Pawlita M, Krammer P . Cytotoxicity-dependent APO-1 (Fas/CD95)-associated proteins form a death-inducing signaling complex (DISC) with the receptor. EMBO J. 1995; 14(22):5579-88. PMC: 394672. DOI: 10.1002/j.1460-2075.1995.tb00245.x. View

12.

Kochan J, Murialdo H . Early intermediates in bacteriophage lambda prohead assembly. II. Identification of biologically active intermediates. Virology. 1983; 131(1):100-15. DOI: 10.1016/0042-6822(83)90537-8. View

13.

Medema J, Scaffidi C, Kischkel F, Shevchenko A, Mann M, Krammer P . FLICE is activated by association with the CD95 death-inducing signaling complex (DISC). EMBO J. 1997; 16(10):2794-804. PMC: 1169888. DOI: 10.1093/emboj/16.10.2794. View

14.

Zhang J, Winoto A . A mouse Fas-associated protein with homology to the human Mort1/FADD protein is essential for Fas-induced apoptosis. Mol Cell Biol. 1996; 16(6):2756-63. PMC: 231266. DOI: 10.1128/MCB.16.6.2756. View

15.

Takashina T, Nakayama M . Modifications enhance the apoptosis-inducing activity of FADD. Mol Cancer Ther. 2007; 6(6):1793-803. DOI: 10.1158/1535-7163.MCT-06-0522. View