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Upstream control of apoptosis by caspase-2 in serum-deprived primary neurons

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Abstract

During development as well as in pathological situations, neurons that fail to find appropriate targets or neurotrophic factors undergo cell death. Using primary cortical neurons subjected to acute serum-deprivation (SD), we have examined caspases activation, mitochondrial dysfunction and cell death parameters. Among a panel of metabolic, signaling and caspases inhibitors only those able to interfere with caspase-2 like activity protect primary neurons against SD-induced cell death. In situ detection and subcellular fractionation demonstrate a very early activation of cytosolic caspase-2, which controls Bax cleavage, relocalization and mitochondrial membrane permeabilization (MMP). Both z-VDVAD-fmk and a siRNA specific for caspase-2 abolish Bax changes, mitochondrial membranes permeabilization, as well as cytochrome c release-dependent activation of caspase-9/caspase-3, nuclear alterations, phosphatidylserine exposure, neurites dismantling and neuronal death. Hence, caspase-2 is an early checkpoint for apoptosis initiation in primary neurons subjected to serum deprivation.

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Abbreviations

7-AAD:

7-Amino Actinomycin D

ALLM:

N- Acetyl-Leu-Leu-Met-al

ALLN:

N-Acetyl-Leu-Leu-Norleu-al

BAPTA-AM:

acetoxymethyl ester of 1, 2-bis(2-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid

BOC-D-FMK:

BOC-Asp (OMe)-fluoromethylketone

mClCCP:

carbonylcyanide m- chlorophenylhydrazone

CMXRos:

Mito

Tracker® Red CMXRos

ΔΨ m :

mitochondrial transmembrane potential

DIDS:

4,4′-Diisothiocyanastilbene-2,2′-disulfonic acid disodium salt

E64d:

trans-Epoxysuccinyl-L-leucylamido-(4- guanidine)butane

FLICA:

Fluorochrome-Labeled Inhibitor of Caspase; FC: flow cytometry

FM:

fluorescence micro- scopy

JC-1:

5,5′,6,6′-tetrachloro-1,1′, 3,3′-tetraethyl- benzimidazolylcarbocyanine iodide

MMP:

mitochondrial membrane permeabilization

MPTP:

1-methyl-4-phenyl-1,2, 3,6-tetrahydropyridine

NA:

nuclear apoptosis

Pefabloc SC®:

4-(2-Aminoethyl)-benzenesulfonyl fluoride

SD:

serum deprivation

PS:

phosphatidyl-serine

PTP:

permeability transition pore

Q-VD-OPH:

Quinoline-Val-Asp (OMe)-CH2-O-Ph, Q-VD-OPH; Z-DEVD-FMK; N-benzyloxycarbonyl-Asp-Glu(Ome)-His-Asp(Ome)-fluoromethylketone

Z-FA-FMK;:

N-ben- zyloxycarbonyl-Phe-Ala-fluoromethylketone

z-FF-fmk:

N- benzyloxycarbonyl-Phe-Phe-fluoromethylketone

Z-LEHD-:

FMK: N-ben-zyloxycarbonyl-Leu-Glu(Ome)-His-Asp(OMe)-fluoromethylketone

Z-LETD-FMK:

N-benzyloxycarbonyl-Leu-Glu(Ome)-Thr-Asp(OMe)-fluoromethylketone

Z-VAD-FMK:

N-benzyloxycarbonyl-Val-Ala-Asp(Ome)-fluoromethylketone

Z-VDVAD-FMK:

N-benzyloxycarbonyl-Val-Asp(Ome)-Val-Ala-Asp(OMe)-fluoromethylketone

VDAC:

voltage-dependent-anionic-channel

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Author notes

  1. D. Rebouillat

    Present address: Theraptosis, Institut Pasteur, Pasteur BioTop, 28 rue du Dr. Roux, F-75724, Paris cedex 15, France

Authors and Affiliations

  1. Theraptosis Research Laboratory, Theraptosis S.A., Pasteur Biotop, Institut Pasteur, 25-28 Rue du Docteur Roux, 75015, Paris, France

    D. Chauvier, H. Lecoeur, A. Langonné, A. Borgne-Sanchez, D. Rebouillat & E. Jacotot

  2. UMR CNRS 7102 and Université Pierre et Marie Curie, 9 quai Saint Bernard, 75005, Paris, France

    J. Mariani

  3. Department of Cell Biology, Sciences III, University of Geneva, Geneva 4, Switzerland

    J.-C. Martinou

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  1. D. Chauvier
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  2. H. Lecoeur
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Correspondence to D. Chauvier.

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D. Rebouillat and E. Jacotot share senior co-authorship.

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Chauvier, D., Lecoeur, H., Langonné, A. et al. Upstream control of apoptosis by caspase-2 in serum-deprived primary neurons. Apoptosis 10, 1243–1259 (2005). https://doi.org/10.1007/s10495-005-1681-x

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