Head-down tilt increases rat cardiac muscle eIF-2 alpha phosphorylation

We previously demonstrated that head-down tilt in rats decreases heart polypeptide initiation rate and proposed a mechanism whereby redistribution of the chaperone heat-shock cognate/heat-shock protein-70 (HSC/HSP-70) facilitates the phosphorylation of eukaryotic initiation factor-2 alpha (eIF-2 alpha). In this study, two-dimensional gel electrophoretic analysis of eIF-2 alpha showed no phosphorylation in control hearts. At 8 h of head-down tilt, there was a 45% increase in total eIF-2 alpha, and 79% was phosphorylated. At 18 h, eIF-2 alpha increased to 142% of control, of which 4% was phosphorylated. This is consistent with the previous study where, at 8 h, there was a 78% increase in polysomal HSC/HSP-70 and a shift in the polysome center-of-mass to lighter polysomes (indicating decreased initiation). After 18 h of suspension, polysomal HSC/HSP-70 levels were 24% relative to control, and the center-of-mass returned toward control. We conclude that the decrease in polypeptide initiation during head-down tilt is mediated by HSC/HSP-70 via phosphorylation of eIF-2 alpha.

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Effect of Brain Ischemia and Reperfusion on the Localization of Phosphorylated Eukaryotic Initiation Factor 2α

Journal of Cerebral Blood Flow & Metabolism ◽

10.1097/00004647-199712000-00004 ◽

1997 ◽

Vol 17 (12) ◽

pp. 1291-1302 ◽

Cited By ~ 92

Author(s):

Donald J. DeGracia ◽

Jonathon M. Sullivan ◽

Robert W. Neumar ◽

Sarah S. Alousi ◽

Katie R. Hikade ◽

...

Keyword(s):

Protein Synthesis ◽

Translation Initiation ◽

Initiation Factor ◽

Eukaryotic Initiation Factor ◽

Initiation Rate ◽

Western Blots ◽

Α Subunit ◽

Neuronal Nuclei ◽

Initiation Factor 2 ◽

Brain Reperfusion

Postischemic brain reperfusion is associated with a substantial and long-lasting reduction of protein synthesis in selectively vulnerable neurons. Because the overall translation initiation rate is typically regulated by altering the phosphorylation of serine 51 on the α-subunit of eukaryotic initiation factor 2 (eIF-2α), we used an antibody specific to phosphorylated eIF-2α [eIF-2(αP)] to study the regional and cellular distribution of eIF-2(αP) in normal, ischemic, and reperfused rat brains. Western blots of brain postmitochondrial supernatants revealed that ~1% of all eIF-2α is phosphorylated in controls, eIF-2(αP) is not reduced by up to 30 minutes of ischemia, and eIF-2(αP) is increased ~20-fold after 10 and 90 minutes of reperfusion. Immunohistochemistry shows localization of eIF-2(αP) to astrocytes in normal brains, a massive increase in eIF-2(αP) in the cytoplasm of neurons within the first 10 minutes of reperfusion, accumulation of eIF-2(αP) in the nuclei of selectively vulnerable neurons after 1 hour of reperfusion, and morphology suggesting pyknosis or apoptosis in neuronal nuclei that continue to display eIF-2(αP) after 4 hours of reperfusion. These observations, together with the fact that eIF-2(αP) inhibits translation initiation, make a compelling case that eIF-2(αP) is responsible for reperfusion-induced inhibition of protein synthesis in vulnerable neurons.

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