Alteration of S-nitrosothiol homeostasis and targets for protein S-nitrosation in human hepatocytes.
The liver is one organ clearly influenced by nitric oxide (NO), and acute and chronic exposure to this substance has been associated with distinct patterns of liver disease. Disruption or deregulation of S-nitrosothiol (SNO) signalling leads to impairment of cellular function and disease, and this study was aimed to identify potential targets for protein S-nitrosation during alteration of SNO homeostasis in human hepatocytes. Cells were treated with S-nitroso-L-cysteine (CSNO), an effective physiological nitrosothiol for delivering NO bioactivity to cells. Treatment with CSNO augmented the levels of S-nitrosoproteins detected both by chemiluminescence and the biotin switch method. CSNO treatment also increased S-nitrosoglutathione reductase (GSNOR) activity that returned SNO content to basal levels. This increased enzymatic activity was related to augmented levels of ADH-5 mRNA, the gene encoding for GSNOR in humans. In addition, the treatment with the SNO also increased cell death. Twenty S-nitrosoproteins were identified in CSNO-treated hepatocytes, including mitochondrial aldehyde dehydrogenase, protein disulphide isomerase, Hsp60, GRP75 and Raf kinase inhibitor protein. The identification in the S-nitrosatable proteome of proteins involved in metabolism, maintenance of cellular homeostasis and signalling points to the relevance of protein S-nitrosation to the physiology and pathophysiology of human hepatocytes.
Liver Research Unit, Hospital Universitario Reina Sofía, Córdoba, Spain.
Laura M
LM
López-Sánchez
Fernando J
FJ
Corrales
Raúl
R
González
Gustavo
G
Ferrín
Juan R
JR
Muñoz-Castañeda
Isidora
I
Ranchal
Ana B
AB
Hidalgo
Javier
J
Briceño
Pedro
P
López-Cillero
Miguel A
MA
Gómez
Manuel
M
De La Mata
Jordi
J
Muntané
Antonio
A
Rodríguez-Ariza
D000328:Adult; D000368:Aged; D000445:Aldehyde Oxidoreductases / Q000235:genetics / Q000378:metabolism; D000704:Analysis of Variance; D001710:Biotin; D002853:Chromatography, Liquid; D003545:Cysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D005260:Female; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism*; D006706:Homeostasis; D006801:Humans; D008107:Liver Diseases / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000378:metabolism*; D015538:Nitrosation; D049488:Phosphatidylethanolamine Binding Protein / Q000378:metabolism; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D026403:S-Nitrosothiols / Q000378:metabolism* / Q000494:pharmacology; D053719:Tandem Mass Spectrometry; D014407:Tumor Cells, Cultured / Q000378:metabolism
D049488:Phosphatidylethanolamine Binding Protein; D011506:Proteins; D026403:S-Nitrosothiols; D009569:Nitric Oxide; D001710:Biotin; C021315:S-nitrosocysteine; D000445:Aldehyde Oxidoreductases; C087006:formaldehyde dehydrogenase, glutathione-independent; D019704:Protein Disulfide-Isomerases; D003545:Cysteine