Functional and structural alterations in the endoplasmic reticulum and mitochondria during apoptosis triggered by C2-ceramide and CD95/APO-1/FAS receptor stimulation.
C(2)-ceramide (C(2)-cer) and binding of the CD95/APO-1/FAS (aCD95) receptor are acknowledged inducers of apoptosis. In spite of that, their effects on the endoplasmic reticulum (ER) and mitochondria during early phases of apoptotic onset are poorly characterized. Here, by employing various approaches, we followed structural and functional modifications of these organelles at the beginning of cellular demise. In detail, we observed that C(2)-cer, but not CD95 activation, markedly modifies the morphology of the ER and promotes Ca(2+) release. Accordingly, mitochondria of C(2)-cer-treated, but not of CD95-stimulated, cells are fragmented, show reduced Ca(2+) uptake, and collapsed membrane potential (DeltaPsi(m)). Most notably, C(2)-cer-mediated morphological aberrations of the ER are prevented neither by the pan-caspase inhibitor Z-VADfmk nor by the cell cytoskeleton dissembler cytochalasin-D, while on the contrary they are reduced by incubation in the presence of the intracellular Ca(2+) chelator 1,2-bis(O-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). We conclude that initiation of apoptosis via the intrinsic (i.e. C(2)-cer mediated) pathway causes an early structural and functional alteration of both ER and mitochondria, thus underlying a final "non return" point in the apoptotic pathway.
Ferrari
Davide
D
Department of Experimental and Diagnostic Medicine, Section of General Pathology, Interdisciplinary Center for the Study of Inflammation and Emilia Romagna Laboratory BioPharmaNet, University of Ferrara, Ferrara, Italy.
Pinton
Paolo
P
Campanella
Michelangelo
M
Callegari
Maria Giulia
MG
Pizzirani
Cinzia
C
Rimessi
Alessandro
A
Di Virgilio
Francesco
F
Pozzan
Tullio
T
Rizzuto
Rosario
R
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism; D061945:Caspase Inhibitors; D002614:Chelating Agents / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure*; D006367:HeLa Cells; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000378:metabolism / Q000648:ultrastructure*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism; D019014:fas Receptor / Q000819:agonists* / Q000378:metabolism
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D002614:Chelating Agents; D015853:Cysteine Proteinase Inhibitors; C064769:N-acetylsphingosine; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D019014:fas Receptor; D004533:Egtazic Acid; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D013110:Sphingosine; D002118:Calcium