ERK1/2 Regulates Intracellular ATP Levels through α-Enolase Expression in Cardiomyocytes Exposed to Ischemic Hypoxia and Reoxygenation

Abstract

Extracellular signal-regulated kinase 1/2 (ERK1/2) is known to function in cell survival in response to various stresses; however, the mechanism of cell survival by ERK1/2 remains poorly elucidated in ischemic heart. Here we applied functional proteomics by two-dimensional electrophoresis to identify a cellular target of ERK1/2 in response to ischemic hypoxia. Approximately 1500 spots were detected by Coomassie Brilliant Blue staining of a sample from unstimulated cells. The staining intensities of at least 50 spots increased at 6-h reoxygenation after 2-h ischemic hypoxia. Of the 50 spots that increased, at least 4 spots were inhibited in the presence of PD98059, a MEK inhibitor. A protein with a molecular mass of 52 kDa that is strongly induced by ERK1/2 activation in response to ischemic hypoxia and reoxygenation was identified as ␣-enolase, a rate-limiting enzyme in the glycolytic pathway, by liquid chromatographymass spectrometry and amino acid sequencing. The expressions of the ␣-enolase mRNA and protein are inhibited during reoxygenation after ischemic hypoxia in the cells containing a dominant negative mutant of MEK1 and treated with a MEK inhibitor, PD98059, leading to a decrease in ATP levels. ␣-Enolase expression is also observed in rat heart subjected to ischemia-reperfusion. The induction of ␣-enolase by ERK1/2 appears to be mediated by c-Myc. The introduction of the ␣-enolase protein into the cells restores ATP levels and prevents cell death during ischemic hypoxia and reoxygenation in these cells. These results show that ␣-enolase expression by ERK1/2 participates in the production of ATP during reoxygenation after ischemic hypoxia, and a decrease in ATP induces apoptotic cell death. Furthermore, ␣-enolase improves the contractility of cardiomyocytes impaired by ischemic hypoxia. Our results reveal that ERK1/2 plays a role in the contractility of cardiomyocytes and cell survival through ␣-enolase expression during ischemic hypoxia and reoxygenation.

Neurodegeneration in Heterozygous Niemann-Pick Type C1 (NPC1) Mouse

“…Previous studies demonstrated that mitochondrial dysfunction with energy depletion causes cell death (33,34) and that tau abnormalities alone can cause neurodegenerative diseases (35,36), indicating that each pathway can independently cause neuronal death. However, previous studies showed that there is a correlation between ATP level and the activity of tau kinases including MAPK/ERK1/2 (37,38). These results enable us to assume that mitochondrial dysfunction with a decreased ATP level and the enhancement of tau phosphorylation synergistically contribute to neurodegeneration in aged NPC1ϩ/Ϫ mouse brains.…”

The Role of ERK1/2 Signaling Pathway in Coronary Microembolization-Induced Rat Myocardial Inflammation and Injury

“…Numerous reports indicate that the ERK1/2 signaling pathway plays an important role in cardiovascular physiology and pathology, such as cardiac hypertrophy [29], myocyte proliferation [30], endothelial cell proliferation [31], and vascular smooth muscle cells proliferation and migration [32,33], especially in myocardial ischemic-reperfusion injury [13,34,35]. However, little is known about the role of the ERK1/2 signaling pathway in CME.…”

“…While some studies show that ERK1/2 signaling provides beneficial, cardioprotective effects on certain cardiac pathological processes, such as ischemia and reperfusion [13,14,15,16,17], growing evidence indicates that ERK1/2 is also involved in cardiovascular inflammation [18,19,20]. However, it remains unknown whether ERK1/2 plays a role in CME.…”

Peptide fractionation in proteomics approaches

“…This separation is characterized by the use of a hydrophilic stationary phase, that is, cyano-, diol-, amino-and other bonded phases [18], and a hydrophobic organic mobile phase [19]. The elution step occurs by increasing the polarity of the mobile phase, normally by performing a water gradient resulting in an order of elution inverted relative to RPLC [19,20], that is, the less polar peptides elute first and the more polar peptides elute later. HILIC has also been found to allow enrichment and the targeted ana lysis of posttranslational modifications, such as glycosylation, N-acetylation and phosphorylation, in proteomics applications [19].…”