Alix, a protein regulating endosomal trafficking, is involved in neuronal death.
Alix/AIP1 is a cytoplasmic protein, which was first characterized as an interactor of ALG-2, a calcium-binding protein necessary for cell death. Alix has also recently been defined as a regulator of the endo-lysosomal system. Here we have used post-mitotic cerebellar neurons to test Alix function in caspase-dependent and -independent cell death. Indeed, these neurons survived when cultured in 25 mm potassium-containing medium but underwent apoptosis soon after the extracellular potassium was lowered to 5 mm. In agreement with other studies, we show that caspases are activated after K+ deprivation, but that inhibition of these proteases, using the pancaspase inhibitor boc-aspartyl(OMe)-fluoromethylketone, has no effect on cell survival. Transfection experiments demonstrated that Alix overexpression is sufficient to induce caspase activation, whereas overexpression of its C-terminal half, Alix-CT, blocks caspase activation and cell death after K+ deprivation. We also define a 12-amino acid PXY repeat of the C-terminal proline-rich domain necessary for binding ALG-2. Deletion of this domain in Alix or in Alix-CT abolished the effects of the overexpressed proteins on neuronal survival, demonstrating that the ALG-2-binding region is crucial for the death-modulating function of Alix. Overall, these findings define the Alix/ALG-2 complex as a regulator of cell death controlling both caspase-dependent and -independent pathways. They also suggest a molecular link between the endo-lysosomal system and the effectors of the cell death machinery.
Laboratoire Neurodégénérescence et Plasticité, INSERM-UJF, Pavillon de Neurologie, Hopital A. Michallon, 38043 Grenoble Cedex 9, France.
Yaël
Y
Trioulier
Sakina
S
Torch
Béatrice
B
Blot
Nadine
N
Cristina
Christine
C
Chatellard-Causse
Jean-Marc
JM
Verna
Rémy
R
Sadoul
D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D002352:Carrier Proteins / Q000502:physiology*; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D011992:Endosomes / Q000378:metabolism*; D051379:Mice; D009474:Neurons / Q000502:physiology*
D051017:Apoptosis Regulatory Proteins; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; C490815:Pdcd6 protein, mouse; C492144:Pdcd6ip protein, mouse