Apoptosis Sherwin Wilk, Ph.D. Mount Sinai School of Medicine Department of Pharmacology and Biological Chemistry Cell Signaling Systems Course Spring 2005

Wojcik, C. et al, Apoptosis. 1997;2(5):

C. elegans Death Genes Pro-apoptotic Pro-apoptotic ced-3 ced-3 ced-4 ced-4 Anti-apoptotic ced-9

Pro-IL-1β (31 – 33 kDa) IL-1-β converting enzyme IL-1-β (17.5 kDa) Single cleavage at Asp 116 -Ala 117

Substrate specificity of ICE P 4 -P 3 -P 2 -P 1 -P 1 ´-P 2 ´-P 3 ´ Asp is required in P 1 Synthetic Substrate acetyl-Tyr-Val-Ala-Asp-amc ac-YVAD-amc

ICE is a cysteine proteinase It is inactivated by –SH blocking reagents It is inactivated by –SH blocking reagents 14 C-iodoacetate is incorporated into the 20 kDa subunit 14 C-iodoacetate is incorporated into the 20 kDa subunit The enzyme can be potently inhibited by a peptide aldehyde The enzyme can be potently inhibited by a peptide aldehyde

Thornberry et al, Nature Apr 30;356(6372):

crmA (cytokine response modifier) 38 kDa cowpox virus proteinase inhibitor 38 kDa cowpox virus proteinase inhibitor Member of the serpin family Member of the serpin family Inhibits ICE Inhibits ICE Inhibits apoptosis Inhibits apoptosis

DNA endonuclease oligonucleotides (-) inhibited by poly ADP ribosylation

Nicholson et al, Nature Jul 6;376(6535): PARP cleavage product

Nicholson et al, Nature Jul 6;376(6535):37-43.

Pro P17P12 ^ QACRG DNDS

Alnemri et al, Cell Oct 18;87(2):171

Stennicke and Salvesen, Biochim Biophys Acta Sep 8;1387(1-2):17-31.

Bcl-2 (B cell lymphoma oncogene), a Ced-9 homolog Large protein family Large protein family Many family members reside in the cytoplasmic face of the mitochondrial membrane Many family members reside in the cytoplasmic face of the mitochondrial membrane Transmits a survival signal when transfected into cells Transmits a survival signal when transfected into cells Prevents cytochrome C release from mitochondria Prevents cytochrome C release from mitochondria

Extrinsic Apoptotic Pathway TNF-related apoptosis-inducing ligand (TRAIL) pathway

Schulze-Osthoff, Trends Cell Biol Dec;4(12):

Tartaglia et al, Cell Sep 10;74(5):

Ashkenazi and Dixit, Science Aug 28;281(5381): Figure 1. Apoptosis signaling by CD95. DD, death domain; DED, death effector domain. Fig. 2. Proapoptotic and antiapoptotic signaling by TNFR1 and DR3.

Comparison of signaling for NF- κ B or for apoptosis NF- κ B TNFR1 TNFR1 TRADD TRADD TRAF RIP TRAF RIP Apoptosis TNFR1 TRADD FADD caspase 8 downstream ICE signaling

Nagata, Cell Feb 7;88(3):

Enari et al, Nature Jan 1;391(6662):43-50.

Intrinsic (mitochondrial) apoptotic pathway

Cell-free system for the activation of CPP32 (caspase 3) Liu et al, Cell Jul 12;86(1): Activation requires: dATP Apaf-1 Apaf-2 (cytochrome C)

Liu et al, Cell Jul 12;86(1):

Zou et al, Cell Aug 8;90(3):

Proteins modulating mitochondrial apoptosis IAP (inhibitors of apoptosis) – directly binds to active caspases IAP (inhibitors of apoptosis) – directly binds to active caspases Smac [Diablo] (second mitochondrial activator of caspase) – directly binds IAP Smac [Diablo] (second mitochondrial activator of caspase) – directly binds IAP AIF (apoptosis – inducing factor) and endonuclease G – involved in DNA fragmentation AIF (apoptosis – inducing factor) and endonuclease G – involved in DNA fragmentation Omi/HtrA2 (a serine proteinase) – interacts with IAP Omi/HtrA2 (a serine proteinase) – interacts with IAP

Finkel, Science Apr 27;292(5517): Center stage in apoptosis· In this view, numerous cell-death stimuli work through the mitochondrion. They cause pro-apoptotic members of the BCL-2 family, such as BAX and BAK, to either open new pores or modify existing channels in the mitochondrial membrane, releasing cytochrome c and other proteins that lead to caspase activation and cell death. BCL-2 itself, which is antiapoptotic, somehow blocks the pore or channel opening. ILLUSTRATION: C. SLAYDEN

Adams and Cory, Science Aug 28;281(5381): Figure 1. Pathways to cell death in C. elegans and mammals. The CED-9/Bcl-2 family integrates positive and negative signals and arbitrates whether apoptosis should occur; activation of CED-4/Apaf-1 commits to apoptosis, and CED-3/caspases mediate the death process. In mammalian cells, the Bcl-2 family rules on signals from diverse cytotoxic stimuli (for example, cytokine deprivation and exposure to glucocorticoids, DNA damage, or staurosporine). However, the signal induced by engagement of the "death receptor" CD95 proceeds primarily through the adaptor FADD, which directly activates caspase-8 and largely bypasses the Bcl-2 family (see text).