Abstract
The regulation of protein synthesis is essential for maintaining cellular homeostasis, especially during stress responses, and its dysregulation could underlie the development of human diseases. The critical step during translation regulation is the phosphorylation of eukaryotic initiation factor 2 alpha (eIF2α). Here we report the identification of a direct kinase of eIF2α, microtubule affinity-regulating kinase 2 (MARK2), which phosphorylates eIF2α in response to proteotoxic stress. The activity of MARK2 was confirmed in the cells lacking the 4 previously known eIF2α kinases. MARK2 itself was found to be a substrate of protein kinase C delta (PKCδ), which serves as a sensor for protein misfolding stress through a dynamic interaction with heat shock protein 90 (HSP90). Both MARK2 and PKCδ are activated via phosphorylation in proteotoxicity-associated neurodegenerative mouse models and in human patients with amyotrophic lateral sclerosis (ALS). These results reveal a PKCδ-MARK2-eIF2α cascade that may play a critical role in cellular proteotoxic stress responses and human diseases.
Highlights
To maintain a state of fitness during stress, cells have evolved exquisite stress response programs that sense potentially harmful situations and make the necessary adaptations at the molecular and cellular levels
The phosphorylation of eukaryotic initiation factor 2 alpha (eIF2α) by microtubule affinity-regulating kinase 2 (MARK2) was completely blocked by a MARK2-specific antibody but not by an immunoglobulin G (IgG) control (Fig 1A, lane 8), confirming that the observed activity for the eIF2α kinase was associated with the MARK2 protein
Consistent with the previous reports of endoplasmic reticulum (ER) stress in the Cu/Zn superoxide dismutase (SOD1) mouse models of amyotrophic lateral sclerosis (ALS) [46,47], the phosphorylation of protein kinase R (PKR)-like ER-resident kinase (PERK), but not that of general control nonderepressible factor 2 kinase (GCN2), was increased in the spinal cord tissues from symptomatic mutant SOD1 mice (S6B Fig), suggesting that both PERK and MARK2 could contribute to the phosphorylation of eIF2α in these mice. These results suggest that the protein kinase C delta (PKCδ)-MARK2-eIF2α signaling is a previously unrecognized pathway implicated in the neurodegenerative mouse models associated with misfolded SOD1 proteins
Summary
To maintain a state of fitness during stress, cells have evolved exquisite stress response programs that sense potentially harmful situations and make the necessary adaptations at the molecular and cellular levels. Stress signaling pathways are frequently mediated by protein kinases and phosphorylation substrates, whose specificity is determined by their interactions with temporal and spatial regulations [1]. Proteins are responsible for most cellular functions, and the maintenance of protein homeostasis is required for the survival of cells, especially under stress conditions. The first step in translation requires eukaryotic initiation factor 2 (eIF2), which is regulated by phosphorylation of serine 51 (51S) of its alpha subunit (eIF2α), with increased phosphorylation resulting in global attenuation of the translation of most transcripts and enhanced translation of select transcripts encoding stress response-related proteins. The phosphorylation of eIF2α is the central step during the integrated stress response, which
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