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Glucose-dependent phosphorylation signaling pathways and crosstalk to mitochondrial respiration in insulin secreting cells.

ABSTRACT: BACKGROUND:Glucose is the main secretagogue of pancreatic beta-cells. Uptake and metabolism of the nutrient stimulates the beta-cell to release the blood glucose lowering hormone insulin. This metabolic activation is associated with a pronounced increase in mitochondrial respiration. Glucose stimulation also initiates a number of signal transduction pathways for the coordinated regulation of multiple biological processes required for insulin secretion. METHODS:Shotgun proteomics including TiO2 enrichment of phosphorylated peptides followed by liquid chromatography tandem mass spectrometry on lysates from glucose-stimulated INS-1E cells was used to identify glucose regulated phosphorylated proteins and signal transduction pathways. Kinase substrate enrichment analysis (KSEA) was applied to identify key regulated kinases and phosphatases. Glucose-induced oxygen consumption was measured using a XF96 Seahorse instrument to reveal cross talk between glucose-regulated kinases and mitochondrial activation. RESULTS:Our kinetic analysis of substrate phosphorylation reveal the molecular mechanism leading to rapid activation of insulin biogenesis, vesicle trafficking, insulin granule exocytosis and cytoskeleton remodeling. Kinase-substrate enrichment identified upstream kinases and phosphatases and time-dependent activity changes during glucose stimulation. Activity trajectories of well-known glucose-regulated kinases and phosphatases are described. In addition, we predict activity changes in a number of kinases including NUAK1, not or only poorly studied in the context of the pancreatic beta-cell. Furthermore, we pharmacologically tested whether signaling pathways predicted by kinase-substrate enrichment analysis affected glucose-dependent acceleration of mitochondrial respiration. We find that phosphoinositide 3-kinase, Ca2+/calmodulin dependent protein kinase and protein kinase C contribute to short-term regulation of energy metabolism. CONCLUSIONS:Our results provide a global view into the regulation of kinases and phosphatases in insulin secreting cells and suggest cross talk between glucose-induced signal transduction and mitochondrial activation.

SUBMITTER: Santo-Domingo J

PROVIDER: S-EPMC6381748 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Glucose-dependent phosphorylation signaling pathways and crosstalk to mitochondrial respiration in insulin secreting cells.

Santo-Domingo Jaime J Galindo Antonio Núñez AN Cominetti Ornella O De Marchi Umberto U Cutillas Pedro P Dayon Loïc L Wiederkehr Andreas A

Cell communication and signaling : CCS 20190220 1

<h4>Background</h4>Glucose is the main secretagogue of pancreatic beta-cells. Uptake and metabolism of the nutrient stimulates the beta-cell to release the blood glucose lowering hormone insulin. This metabolic activation is associated with a pronounced increase in mitochondrial respiration. Glucose stimulation also initiates a number of signal transduction pathways for the coordinated regulation of multiple biological processes required for insulin secretion.<h4>Methods</h4>Shotgun proteomics i ...[more]

PMID: 30786936

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Project description:BackgroundCellular response to external stimuli requires propagation of corresponding signals through molecular signaling pathways. However, signaling pathways are not isolated information highways, but rather interact in a number of ways forming sophisticated signaling networks. Since defects in signaling pathways are associated with many serious diseases, understanding of the crosstalk between them is fundamental for designing molecularly targeted therapy. Unfortunately, we still lack technology that would allow high throughput detailed measurement of activity of individual signaling molecules and their interactions. This necessitates developing methods to prioritize selection of the molecules such that measuring their activity would be most informative for understanding the crosstalk. Furthermore, absence of the reaction coefficients necessary for detailed modeling of signal propagation raises the question whether simple parameter-free models could provide useful information about such pathways.ResultsWe study the combined signaling network of three major pro-survival signaling pathways: Epidermal Growth Factor Receptor (EGFR), Insulin-like Growth Factor-1 Receptor (IGF-1R), and Insulin Receptor (IR). Our study involves static analysis and dynamic modeling of this network, as well as an experimental verification of the model by measuring the response of selected signaling molecules to differential stimulation of EGF, IGF and insulin receptors. We introduced two novel measures of the importance of a node in the context of such crosstalk. Based on these measures several molecules, namely Erk1/2, Akt1, Jnk, p70S6K, were selected for monitoring in the network simulation and for experimental studies. Our simulation method relies on the Boolean network model combined with stochastic propagation of the signal. Most (although not all) trends suggested by the simulations have been confirmed by experiments.ConclusionThe simple model implemented in this paper provides a valuable first step in modeling signaling networks. However, to obtain a fully predictive model, a more detailed knowledge regarding parameters of individual interactions might be necessary.

Dataset Information

Glucose-dependent phosphorylation signaling pathways and crosstalk to mitochondrial respiration in insulin secreting cells.

Publications

Glucose-dependent phosphorylation signaling pathways and crosstalk to mitochondrial respiration in insulin secreting cells.

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