Glucose-stimulated Protein Synthesis in Pancreatic β-Cells Parallels an Increase in the Availability of the Translational Ternary Complex (eIF2-GTP·Met-tRNAi) and the Dephosphorylation of eIF2α

Gomez, Edith; Powell, M. J. D.; Greenman, Isabel C.; Herbert, Terence P.

doi:10.1074/jbc.m408682200

Cited by 47 publications

(50 citation statements)

References 55 publications

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“…An mTOR independency of eIF2α (de)phosphorylation has been noticed in other cells [36,37]. The glibenclamideinduced dephosphorylation of eEF2 was almost completely prevented by the PKA inhibitor, indicating the existence of an additional way in which PKA stimulates translation.…”

Section: Discussionmentioning

confidence: 68%

Glibenclamide activates translation in rat pancreatic beta cells through calcium-dependent mTOR, PKA and MEK signalling pathways

et al. 2008

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Aims/hypothesis Prolonged exposure of rat beta cells to the insulin secretagogue glibenclamide has been found to induce a sustained increase in basal insulin synthesis. This effect was calcium-dependent and localised in cells that had been degranulated by the drug. Since it was blocked by the translation inhibitor cycloheximide, we examined whether sustained exposure to glibenclamide activates translational factors by calcium-dependent signalling pathways. Methods Purified rat beta cells were cultured with and without glibenclamide in the presence or absence of inhibitors of calcium-dependent signalling pathways before measurement of basal and stimulated protein and insulin synthesis, and assessment of abundance of (phosphorylated) translation factors.Results A 24 h exposure to glibenclamide induced activation of four translation factors, i.e. phosphorylation of eukaryotic initiation factor (eIF) 4e binding protein 1 and ribosomal protein S6 (rpS6), and dephosphorylation of eIF-2α and eukaryotic elongation factor 2. The rise in phospho-rpS6 intensity was localised to a subpopulation of beta cells with low insulin content. This activation of translational factors and the associated elevation of insulin synthesis were completely blocked by the calcium channel blocker verapamil and partially blocked by the mammalian target of rapamycin (mTOR) inhibitor rapamycin, the protein kinase A (PKA) inhibitor Rp-8-Br-cAMPs and the mitogen-activated protein kinase/ extracellular signal-regulated kinase kinase (MEK) inhibitor U0126; a combination of inhibitors exhibited additive effects. Conclusions/interpretation Prolonged exposure to glibenclamide activates protein translation in pancreatic beta cells through the calcium-regulated mTOR, PKA and MEK signalling pathways. The observed intercellular differences in translation activation are proposed as underlying mechanism for functional heterogeneity in the pancreatic beta cell population.

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Section: Discussionmentioning

confidence: 68%

Glibenclamide activates translation in rat pancreatic beta cells through calcium-dependent mTOR, PKA and MEK signalling pathways

et al. 2008

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“…Since it has been shown that glucose starvation can have a suppressive effect on translation under normoxic conditions in certain cell lines (Gomez et al 2004;Vander Mierde et al 2007) we wanted to determine if the translational repression that we observe under hypoxic conditions is merely a consequence of glucose depletion. Our data clearly show that short-term (20 h) glucose depletion under normoxic conditions has little impact on protein synthesis while dramatically inhibiting translation under hypoxic conditions.…”

Section: Discussionmentioning

confidence: 99%

Translational repression during chronic hypoxia is dependent on glucose levels

Thomas¹,

Dias²,

Johannes³

2008

RNA

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Translation is often repressed in cell lines that are exposed to hypoxic conditions (0.5% -1.5% O 2 ) but this repression requires prolonged exposure (> 16 h). We report here that prolonged exposure to hypoxia results in the depletion of glucose from the media and that the loss of glucose correlates with the shut down in translation. Furthermore, we show that the addition of glucose or reoxygenation restores translation in hypoxic PC3 cells. This indicates that both glucose depletion and hypoxia are required for translational repression. We also show that eIF2a phosphorylation is reversed by glucose addition. Moreover, we present data that strongly indicate that eIF2a phosphorylation as well as the translational inhibition that occurs when cells are grown under conditions of glucose depletion and hypoxia is pancreatic eIF2a kinase (PERK) independent. We believe this is the first report to show that glucose depletion is required for translational repression under hypoxic conditions and that this explains why prolonged exposure to hypoxia is required for this inhibition. Since the physiological conditions that lead to tumor hypoxia would also likely lead to reduced glucose levels, understanding the interplay of glucose and hypoxia in regulating tumor metabolism will provide important information on the growth and development of solid tumors.

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“…This indicates that glucose stimulates the recruitment of ribosomes on to mRNAs and hence stimulates an increase in the rate of initiation. Indeed, we have recently shown that glucose stimulates an increase in the initiation step of protein synthesis in MIN6 cells [19]. At low glucose concentration, proinsulin mRNA was found to cosediment with either monosomes or small polysomes (Figures 4ai and 4ii).…”

Section: Glucose Stimulates the Recruitment Of Ribosomes On To Ribosomentioning

confidence: 99%

“…Anti-SRP54 antibody was purchased from Becton Dickinson (Cowley, Oxford, U.K.). Western blotting, SDS/PAGE and immunoprecipitations were performed as described previously [19].…”

Section: Sds/page Western Blotting and Immunoprecipitationmentioning

confidence: 99%

The selective recruitment of mRNA to the ER and an increase in initiation are important for glucose-stimulated proinsulin synthesis in pancreatic β-cells

Greenman

Gomez

Moore

et al. 2005

Biochemical Journal

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Glucose acutely stimulates proinsulin synthesis in pancreatic β-cells through a poorly understood post-transcriptional mechanism. In the present study, we demonstrate in pancreatic β-cells that glucose stimulates the recruitment of ribosomeassociated proinsulin mRNA, located in the cytoplasm, to the ER (endoplasmic reticulum), the site of proinsulin synthesis, and that this plays an important role in glucose-stimulated proinsulin synthesis. Interestingly, glucose has greater stimulatory effect on the recruitment of proinsulin mRNA to the ER compared with other mRNAs encoding secretory proteins. This, as far as we are aware, is the first example whereby mRNAs encoding secretory proteins are selectively recruited to the ER and provides a novel regulatory mechanism for secretory protein synthesis. Contrary to previous reports, and importantly in understanding the mechanism by which glucose stimulates proinsulin synthesis, we demonstrate that there is no large pool of 'free' proinsulin mRNA in the cytoplasm and that glucose does not increase the rate of de novo initiation on the proinsulin mRNA. However, we show that glucose does stimulate the rate of ribosome recruitment on to ribosome-associated proinsulin mRNA. In conclusion, our results provide evidence that the selective recruitment of proinsulin mRNA to the ER, together with increases in the rate of initiation are important mediators of glucose-stimulated proinsulin synthesis in pancreatic β-cells.

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Glucose-stimulated Protein Synthesis in Pancreatic β-Cells Parallels an Increase in the Availability of the Translational Ternary Complex (eIF2-GTP·Met-tRNAi) and the Dephosphorylation of eIF2α

Cited by 47 publications

References 55 publications

Glibenclamide activates translation in rat pancreatic beta cells through calcium-dependent mTOR, PKA and MEK signalling pathways

Glibenclamide activates translation in rat pancreatic beta cells through calcium-dependent mTOR, PKA and MEK signalling pathways

Translational repression during chronic hypoxia is dependent on glucose levels

The selective recruitment of mRNA to the ER and an increase in initiation are important for glucose-stimulated proinsulin synthesis in pancreatic β-cells

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