Abstract
The anti-apoptotic molecule Aven was originally identified in a yeast two-hybrid screen for Bcl-xL-interacting proteins and has also been found to bind Apaf-1, thereby interfering with Apaf-1 self-association during apoptosome assembly. Aven is expressed in a wide variety of adult tissues and cell lines, and there is increasing evidence that its overexpression correlates with tumorigenesis, particularly in acute leukemias. The mechanism by which the anti-apoptotic activity of Aven is regulated remains poorly understood. Here we shed light on this issue by demonstrating that proteolytic removal of an inhibitory N-terminal Aven domain is necessary to activate the anti-apoptotic potential of the molecule. Furthermore, we identify Cathepsin D (CathD) as the protease responsible for Aven cleavage. On the basis of our results, we propose a model of Aven activation by which its N-terminal inhibitory domain is removed by CathD-mediated proteolysis, thereby unleashing its cytoprotective function.
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Abbreviations
- Ac-DEVD-AFC:
-
N-acetyl-Asp-Glu-Val-Asp-7-amino-4-trifluoromethylcoumarin
- Apaf-1:
-
apoptotic protease activating factor 1
- ATM:
-
ataxia telangiectasia mutated
- Bcl-xL:
-
B cell lymphoma extra large
- Bcl-2:
-
B-cell lymphoma 2
- Bid:
-
BH3 interacting domain death agonist
- CathD:
-
Cathepsin D
- CED-4:
-
cell death abnormality 4
- Cyt c:
-
cytochrome c
- dATP:
-
desoxy-adenosine-triphosphate
- DMEM:
-
Dulbecco's modified Eagle's medium
- DMSO:
-
dimethyl sulfoxide
- DTT:
-
dithiothreitol
- EDTA:
-
ethylenediaminetetraacetic acid
- E64:
-
N-[N-(L-3-trans-carboxyirane-2-carbonyl)-L-leucyl]-agmatine
- FasL:
-
Fas ligand
- FBS:
-
fetal bovine serum
- GFP:
-
green flourescent protein
- IP:
-
immunoprecipitation
- LAV:
-
large intracellular acidic vesicles
- MES:
-
2-(N-morpholino)ethanesulfonic acid
- NaCl:
-
sodium chloride
- NHS AcD3-acetate:
-
N-hydroxysuccinimide trideutero-acetate
- PEI:
-
polyethylenimine
- pAPMSF:
-
p-amidino-phenylmethylsulfonylfluoride
- ProT:
-
prothymosin alpha
- RPMI:
-
1640 Roswell Park Memorial Institute
- TNF:
-
tumor necrosis factor
- z-VAD:
-
carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-luoromethylketone
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Acknowledgements
We thank R Jänicke (University of Düsseldorf, Germany), M Rimpler (MDC Berlin, Germany), GM Cohen (MRC Toxicology Unit, University of Leicester, UK), R Eskes (Molecular Hematology, University Clinic Frankfurt, Germany), and JM Hardwick (John Hopkins University, Baltimore, USA) for generously supplying the cells, expression constructs and antiserum (see Materials and Methods for details). FI is a Postdoctoral Fellow of the Research Foundation – Flanders (FWO-Vlaanderen). This work was supported by grants from the German Cancer Aid Foundation (no. 108659; MZ) and the German National Genome Research Network (NGFN project N1KR-S12T23, MZ). Furthermore, the financial support by the Ministerium für Innovation, Wissenschaft und Forschung des Landes Nordrhein-Westfalen is gratefully acknowledged. KG acknowledges support from a research project of the FWO-Vlaanderen (G.0048.08).
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Melzer, I., Fernández, S., Bösser, S. et al. The Apaf-1-binding protein Aven is cleaved by Cathepsin D to unleash its anti-apoptotic potential. Cell Death Differ 19, 1435–1445 (2012). https://doi.org/10.1038/cdd.2012.17
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DOI: https://doi.org/10.1038/cdd.2012.17
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