Implication of eIF2α kinase GCN2 in induction of apoptosis and endoplasmic reticulum stress-responsive genes by sodium salicylate.
OBJECTIVE
Sodium salicylate (NaSal) can disturb cell viability by affecting the activity of multiple cellular molecules. In this work, we investigated the involvement of stress-responsive kinase GCN2 in regulating cell death and expression of stress genes in mouse embryonic fibroblasts (MEFs) upon exposure to NaSal.


METHODS
Cell viability was assayed using the 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) method, and apoptosis was evaluated by annexin V and propidium iodide staining. A polymerase chain reaction (PCR) array approach was used to analyse differential expression of a panel of 84 endoplasmic reticulum (ER) stress-associated genes. Gene reporter assays were carried out to determine activity of ER stress element (ERSE), and the protein levels of activating transcription factor 6 (ATF6) and C/EBP homologous protein (CHOP) were determined by western blot.


RESULTS
NaSal treatment resulted in reduction of cellular viability and induction of apoptosis in wild-type but not Gcn2(-/-)  cells. Many genes with important functions in protein synthesis/degradation, transcriptional regulation and apoptosis were induced by NaSal and most of these were dependent on GCN2. The activation of ERSE within Ddit3 and the production of CHOP and ATF6 induced by NaSal required GCN2.


CONCLUSIONS
Our data provide evidence for the involvement of GCN2 in apoptosis and gene expression triggered by NaSal, and contributes to the understanding of molecular events occurring in NaSal-treated cells.
Laboratory of Inflammatory Genes, Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, Brazil.
Solange H L
SH
Gentz
Caryne M
CM
Bertollo
Elaine M
EM
Souza-Fagundes
Aristóbolo Mendes
AM
da Silva
D051702:Activating Transcription Factor 6 / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D059865:Endoplasmic Reticulum Stress / Q000235:genetics*; D005347:Fibroblasts / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D051379:Mice; D014176:Protein Biosynthesis / Q000187:drug effects / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D012980:Sodium Salicylate / Q000494:pharmacology*; D051742:Transcription Factor CHOP / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics
D051702:Activating Transcription Factor 6; C497112:Atf6 protein, mouse; C497250:Ddit3 protein, mouse; D051742:Transcription Factor CHOP; C070634:Eif2ak4 protein, mouse; D017346:Protein Serine-Threonine Kinases; D012980:Sodium Salicylate