Phosphorylation of Eukaryotic Initiation Factor 2 by Heme-Regulated Inhibitor Kinase-Related Protein Kinases in <i>Schizosaccharomyces pombe</i> Is Important for Resistance to Environmental Stresses

Zhan, Ke; Vattem, Krishna M.; Bauer, Bettina N.; Dever, Thomas E.; Chen, Jane-Jane; Wek, Ronald C.

doi:10.1128/mcb.22.20.7134-7146.2002

Cited by 71 publications

(60 citation statements)

References 91 publications

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“…In particular, His-119 and Cys-409, the proposed heme axial ligands, are conserved only in mammals. Because the HRI homolog from Schizosaccharomyces pombe is not controlled by the heme iron (24), it is assumed that the heme-regulated HRI function is specific to mammals. In addition, erythroid differentiation in mammals is different from that in other vertebrates.…”

Section: Discussionmentioning

confidence: 99%

Elucidation of the Heme Binding Site of Heme-regulated Eukaryotic Initiation Factor 2α Kinase and the Role of the Regulatory Motif in Heme Sensing by Spectroscopic and Catalytic Studies of Mutant Proteins

Igarashi

Murase

Iizuka

et al. 2008

Journal of Biological Chemistry

141

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Heme-regulated eukaryotic initiation factor 2␣ (eIF2␣) kinase (HRI) functions in response to the heme iron concentration. At the appropriate heme iron concentrations under normal conditions, HRI function is suppressed by binding of the heme iron. Conversely, upon heme iron shortage, HRI autophosphorylates and subsequently phosphorylates the substrate, eIF2␣, leading to the termination of protein synthesis. The molecular mechanism of heme sensing by HRI, including identification of the specific binding site, remains to be established. In the present study we demonstrate that His-119/His-120 and Cys-409 are the axial ligands for the Fe(III)-protoporphyrin IX complex (hemin) in HRI, based on spectral data on site-directed mutant proteins. Cys-409 is part of the heme-regulatory CysPro motif in the kinase domain. A P410A full-length mutant protein displayed loss of heme iron affinity. Surprisingly, inhibitory effects of the heme iron on catalysis and changes in the heme dissociation rate constants in full-length His-119/His-120 and Cys-409 mutant proteins were marginally different to wild type. In contrast, heme-induced inhibition of Cys-409 mutants of the isolated kinase domain and N-terminal-truncated proteins was substantially weaker than that of the full-length enzyme. A pulldown assay disclosed heme-dependent interactions between the N-terminal and kinase domains. Accordingly, we propose that heme regulation is induced by interactions between heme and the catalytic domain in conjunction with global tertiary structural changes at the N-terminal domain that accompany heme coordination and not merely by coordination of the heme iron with amino acids on the protein surface.Eukaryotic cells decrease their overall rates of protein synthesis for survival in response to a variety of stress conditions, such as shortage of amino acids, UV light illumination, virus infection, and accumulation of denatured proteins. Much of the decrease in protein synthesis is caused by phosphorylation of eukaryotic initiation factor 2␣ (eIF2␣) 4 at Ser-51 by eIF2␣ kinases that respond specifically to stress (1-4). Heme-regulated eIF2␣ kinase or heme-regulated inhibitor (HRI) is a member of the eIF2␣ kinase family that controls globin synthesis in response to the heme concentration in reticulocytes (5-8). HRI is inactive at normal heme concentrations. Under conditions of heme deficiency, the enzyme is activated by autophosphorylation and subsequently phosphorylates eIF2␣ at Ser-51. In addition to globin, HRI controls the synthesis of tryptophan 2,3-dioxygenase and cytochrome P450 2B in liver upon acute porphyria (9, 10). Thus, HRI is possibly one of the most important existing heme sensor protein families for eukaryote survival in response to cell emergency states.Heme-responsive/sensing proteins, also known as "heme sensor proteins" are a current focus of investigation. In these proteins heme association (or dissociation) per se regulates various important physiological functions, such as transcription, proteolysis, and kinase activity (11...

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Section: Discussionmentioning

confidence: 99%

Elucidation of the Heme Binding Site of Heme-regulated Eukaryotic Initiation Factor 2α Kinase and the Role of the Regulatory Motif in Heme Sensing by Spectroscopic and Catalytic Studies of Mutant Proteins

Igarashi

Murase

Iizuka

et al. 2008

Journal of Biological Chemistry

141

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“…As shown in S. cerevisiae this post-translational event occurs at the onset of the stress response in a Gcn2/Ppk28-dependent manner (Mascarenhas et al, 2008). Another eIF2a kinase, Hri2, becomes also critical for survival upon long exposure of fission yeast cells to stress (Dunand-Sauthier et al, 2005;Zhan et al, 2002). Stress-induced activation of the Sty1 pathway triggers translation re-initiation by negatively regulating the closely related pathways of the Gcn2 and Hri2 kinases (Berlanga et al, 2010;Dunand-Sauthier et al, 2005).…”

Section: The Sty1 Regulatory Signals On Translationmentioning

confidence: 99%

Oxidative stress response pathways: Fission yeast as archetype

Papadakis¹,

Workman²

2015

Critical Reviews in Microbiology

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Schizosaccharomyces pombe is a popular model eukaryotic organism to study diverse aspects of mammalian biology, including responses to cellular stress triggered by redox imbalances within its compartments. The review considers the current knowledge on the signaling pathways that govern the transcriptional response of fission yeast cells to elevated levels of hydrogen peroxide. Particular attention is paid to the mechanisms that yeast cells employ to promote cell survival in conditions of intermediate and acute oxidative stress. The role of the Sty1/Spc1/Phh1 mitogen-activated protein kinase in regulating gene expression at multiple levels is discussed in detail.

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“…2A). S. pombe possess, in addition to Gcn2, two other eIF2␣ kinases, Hri1 and Hri2 (33). We expressed GFP-YpkA in strains deleted for either of these kinases and found that YpkA sensitivity in hri1⌬ cells was comparable with wild-type cells (data not shown), whereas hri2⌬ cells were, like cells expressing eIF2␣(S52A), especially sensitive to YpkA (Fig.…”

Section: Resultsmentioning

confidence: 99%

The Activities of the Yersinia Protein Kinase A (YpkA) and Outer Protein J (YopJ) Virulence Factors Converge on an eIF2α Kinase

Wiley

Shrestha

Yang

et al. 2009

Journal of Biological Chemistry

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The Yersinia protein kinase A (YpkA) and outer protein J (YopJ) are co-expressed from a single transcript and are injected directly into eukaryotic cells by the plague bacterium Yersinia pestis. When overexpressed in vertebrate or yeast cells, YpkA disrupts the actin-based cytoskeletal system by an unknown mechanism, whereas YopJ obstructs inductive chemokine expression by inhibiting MAPK and NF-B signaling. Previously, we showed that the fission yeast Schizosaccharomyces pombe was sensitive to the kinase activity of YpkA. Here, we screened yeast for cellular processes important for YpkA activity and found that the eIF2␣ kinases mollify the toxicity imparted by the kinase activity of YpkA. Specifically, strains lacking the eIF2␣ kinase Hri2 were particularly sensitive to YpkA. Unexpectedly, the activity of YopJ, which conferred a phenotype consistent with its inhibitory effect on MAPK signaling, was also found to be dependent on Hri2. When expressed in S. pombe, YopJ sensitized cells to osmotic and oxidative stresses through a Hri2-dependent mechanism. However, when co-expressed with YpkA, YopJ protected cells from YpkA-mediated toxicity, and this protection was entirely dependent on Hri2. In contrast, YopJ did not confer protection against the toxic effects of the Yersinia virulence factor YopE. These findings are the first to functionally link YpkA and YopJ and suggest that eIF2␣ kinases, which are critically important in antiviral defenses and protection against environmental stresses, also play a role in bacterial virulence.The plague bacterium Yersinia pestis, as well as the closely related enteric pathogens Yersinia pseudotuberculosis and Yersinia enterocolitica, are relatively resistant to the antimicrobial killing systems of innate immune cells. This property is largely dependent on a membrane-bound type 3 secretion system (T3SS) 2 that injects proteins (here referred to as "effectors") directly into the eukaryotic cell (1, 2). This study is focused on two such effectors: YpkA and YopJ (YopO and YopP in Y. enterocolitica). Although they are encoded by a single transcription unit on the 70-kb extrachromosomal virulence plasmid of Yersinia, these two effectors shape the bacterial host relationship in very different ways. YpkA and its associated kinase activity is necessary for the immediate survival of Yersinia following attachment to host cells; this was suggested by animal infection experiments and confirmed at the cellular level in a function-based "infectivity" assay that measures both survival and growth of macrophage-associated Yersinia (3, 4). In contrast, there are no discernible differences in the infectivity assay between the wild-type and ⌬yopJ mutant strains (5). However, YopJ very efficiently blocks MAPK-and NF-B-mediated signaling pathways and the resulting inductive expression of proinflammatory chemokines; the consequences of YopJ activity during an actual infection is a reduction in the local inflammatory response (6 -10). Therefore, unlike YpkA, which is important for the immediate surviv...

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Phosphorylation of Eukaryotic Initiation Factor 2 by Heme-Regulated Inhibitor Kinase-Related Protein Kinases in Schizosaccharomyces pombe Is Important for Resistance to Environmental Stresses

Cited by 71 publications

References 91 publications

Elucidation of the Heme Binding Site of Heme-regulated Eukaryotic Initiation Factor 2α Kinase and the Role of the Regulatory Motif in Heme Sensing by Spectroscopic and Catalytic Studies of Mutant Proteins

Elucidation of the Heme Binding Site of Heme-regulated Eukaryotic Initiation Factor 2α Kinase and the Role of the Regulatory Motif in Heme Sensing by Spectroscopic and Catalytic Studies of Mutant Proteins

Oxidative stress response pathways: Fission yeast as archetype

The Activities of the Yersinia Protein Kinase A (YpkA) and Outer Protein J (YopJ) Virulence Factors Converge on an eIF2α Kinase

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