Changes in Protein Expression and Lysine Acetylation Induced by Decreased Glutathione Levels in Astrocytes.
Astrocytes and neurons form a highly specialized functional unit, and the loss or gain of astrocytic functions can influence the initiation and progression of different neurodegenerative diseases. Neurons depend on the antioxidant protection provided by neighboring astrocytes. Glutathione (γ-l-glutamyl-l-cysteinyl-glycine) is a major component of the antioxidant system that defends cells against the toxic effects of reactive oxygen/nitrogen species. A decline in glutathione levels has been observed in aging and neurodegenerative diseases, and it aggravates the pathology in an amyotrophic lateral sclerosis-mouse model. Using a SILAC-based quantitative proteomic approach, we analyzed changes in global protein expression and lysine acetylation in primary astrocyte cultures obtained from wild-type mice or those deficient in the glutamate-cysteine ligase modifier subunit (GCLM). GCLM knockout astrocytes display an ∼80% reduction in total glutathione levels. We identified potential molecular targets and novel sites of acetylation that are affected by the chronic decrease in glutathione levels and observed a response mediated by Nrf2 activation. In addition, sequence analysis of peptides displaying increased acetylation in GCLM knockout astrocytes revealed an enrichment of cysteine residues in the vicinity of the acetylation site, which suggests potential crosstalk between lysine-acetylation and cysteine modification. Regulation of several metabolic and antioxidant pathways was observed at the level of protein expression and lysine acetylation, revealing a coordinated response involving transcriptional and posttranslational regulation.
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425. vargasm@musc.edu pehar@musc.edu.
Mariana
M
Pehar
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425.
Lauren E
LE
Ball
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425.
Deep R
DR
Sharma
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425.
Benjamin A
BA
Harlan
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425.
Susana
S
Comte-Walters
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425.
Benjamin A
BA
Neely
From the ¶Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425. vargasm@musc.edu pehar@musc.edu.
Marcelo R
MR
Vargas
D000107:Acetylation; D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000818:Animals; D001253:Astrocytes / Q000378:metabolism* / Q000473:pathology; D004195:Disease Models, Animal; D005786:Gene Expression Regulation / Q000235:genetics; D005721:Glutamate-Cysteine Ligase / Q000235:genetics; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009474:Neurons / Q000378:metabolism / Q000473:pathology; D014176:Protein Biosynthesis / Q000235:genetics*; D011499:Protein Processing, Post-Translational / Q000235:genetics; D040901:Proteomics; D017382:Reactive Oxygen Species / Q000378:metabolism
D017382:Reactive Oxygen Species; C560749:GCLM protein, mouse; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D008239:Lysine