Pharmaceutical Sciences Faculty Publications

Proteome Analysis of DNA Damage-Induced Neuronal Death Using High Throughput Mass Spectrometry

Document Type

Article

Publication Date

6-18-2004

Journal Title

The Journal of Biological Chemistry

ISSN

0021-9258

Volume

279

Issue

25

First Page

26685

Last Page

26697

DOI

10.1074/jbc.M401274200

PubMed ID

15060066

Abstract

Isotope-coded affinity tag reagents and high throughput mass spectrometry were used to quantitate changes in the expression of 150 proteins in mouse wild-type (p53(+/+)) cortical neurons undergoing DNA damage-induced death. Immunological techniques confirmed several of the changes in protein expression, but microarray analysis indicated that many of these changes were not accompanied by altered mRNA expression. Proteome analysis revealed perturbations in mitochondrial function, free radical production, and neuritogenesis that were not observed in p53-deficient neurons. Changes in Tau, cofilin, and other proteins recapitulated abnormalities observed in neurodegenerative states in vivo. Additionally, DNA damage caused a p53-dependent decrease in expression of members of the protein kinase A (PKA) signaling pathway. PKA inhibition promoted death in the absence of DNA damage, revealing a novel mechanism by which endogenous down-regulation of PKA signaling may contribute to p53-dependent neuronal death. These data demonstrate the power of high throughput mass spectrometry for quantitative analysis of the neuronal proteome.

Keywords

Apoptosis, blotting, western, coloring agents, DNA, complementary, down-regulation, free radicals, genes, immunoblotting, immunohistochemistry, mass spectrometry, transgenic, neurons, peptides, phalloidine, proteome, proteomics, RNA, messenger, signal transduction, transfection, tumor suppressor protein p53

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