Substitution of methionine 35 inhibits apoptotic effects of Abeta(31-35) and Abeta(25-35) fragments of amyloid-beta protein in PC12 cells.
BACKGROUND
Amyloid-beta peptide (AbetaP), the central constituent of senile plaques in Alzheimer's disease (AD) brain, has been shown to be toxic to neuronal cells and that this toxicity is responsible for the progressive cognitive decline associated with this neurodegenerative disease. The precise mechanism of AbetaP action remains to be determined; however, it has been reported that the methionine residue at position 35 plays a pivotal role in the toxicity of the peptide. With this in mind, the present study examines the effect of mutating the methionine to norleucine in the fragments (31-35) and (25-35) of AbetaP, which have methionine at the C-terminal, in order to investigate the influence of this residue on Abeta-mediated toxic effects on PC12 cells.


METHODS
The toxic and apoptotic effects (release of Cyt c, caspase activation, and DNA fragmentation) exerted by the Abeta(31-35) and Abeta(25-35) wild-type peptides and Abeta(31-35)Met-->Nle, Abeta(25-35)Met-->Nle peptides where methionine was substituted with norleucine were investigated on PC12 cells.


RESULTS
The results obtained shown that both peptides induce neurotoxicity in PC12 cells via an apoptotic cell death pathway, including cytochrome c release, caspase activation, and DNA fragmentation. Furthermore, this study reveals that these events were completely abrogated in cells exposed to Abeta peptides in which methionine 35 was substituted by a norleucine residue.


CONCLUSIONS
On the basis of the results obtained in this study, an additional hypothesis involving the amyloid-beta peptide and the role of Met-35 has been proposed to clarify the mechanisms responsible of neurodegeneration in Alzheimer's disease.
Clementi
Maria Elisabetta
ME
Institute Chimica del Riconoscimento Molecolare (CNR), c/o Institute of Biochemistry and Clinical Biochemistry, Faculty of Medicine, Catholic University, Rome, Italy. elisabetta.clementi@icrm.cnr.it
Misiti
Francesco
F
D019943:Amino Acid Substitution / Q000235:genetics; D016229:Amyloid beta-Peptides / Q000737:chemistry / Q000235:genetics* / Q000633:toxicity*; D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000032:analysis; D045304:Cytochromes c / Q000032:analysis; D006801:Humans; D008715:Methionine / Q000737:chemistry* / Q000235:genetics; D009474:Neurons / Q000187:drug effects / Q000201:enzymology; D009646:Norleucine / Q000737:chemistry / Q000235:genetics; D016716:PC12 Cells; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics* / Q000633:toxicity*; D051381:Rats
D016229:Amyloid beta-Peptides; D010446:Peptide Fragments; C074636:amyloid beta-protein (25-35); C113042:amyloid beta-protein (31-35); D009646:Norleucine; D045304:Cytochromes c; D008715:Methionine; D020169:Caspases