Abstract
Background and MethodologyPancreatic beta cells show intercellular differences in their metabolic glucose sensitivity and associated activation of insulin production. To identify protein markers for these variations in functional glucose sensitivity, rat beta cell subpopulations were flow-sorted for their level of glucose-induced NAD(P)H and their proteomes were quantified by label-free data independent alternate scanning LC-MS. Beta cell-selective proteins were also identified through comparison with rat brain and liver tissue and with purified islet alpha cells, after geometrical normalization using 6 stably expressed reference proteins.Principal FindingsAll tissues combined, 943 proteins were reliably quantified. In beta cells, 93 out of 467 quantifiable proteins were uniquely detected in this cell type; several other proteins presented a high molar abundance in beta cells. The proteome of the beta cell subpopulation with high metabolic and biosynthetic responsiveness to 7.5 mM glucose was characterized by (i) an on average 50% higher expression of protein biosynthesis regulators such as 40S and 60S ribosomal constituents, NADPH-dependent protein folding factors and translation elongation factors; (ii) 50% higher levels of enzymes involved in glycolysis and in the cytosolic arm of the malate/aspartate-NADH-shuttle. No differences were noticed in mitochondrial enzymes of the Krebs cycle, beta-oxidation or respiratory chain.ConclusionsQuantification of subtle variations in the proteome using alternate scanning LC-MS shows that beta cell metabolic glucose responsiveness is mostly associated with higher levels of glycolytic but not of mitochondrial enzymes.
Highlights
Insulin-producing beta cells are the body’s central glucose sensors
Quantification of subtle variations in the proteome using alternate scanning LC-mass spectrometry (MS) shows that beta cell metabolic glucose responsiveness is mostly associated with higher levels of glycolytic but not of mitochondrial enzymes
Intercellular variations in glucokinase abundance and activity give rise to proportionate variations in concentrationresponse curves for glucose-induced NAD(P)H, insulin synthesis and secretion [2,4,5,6,7]. This functional heterogeneity is subject to regulation in vivo – as shown in animals exposed to sulfonylurea [8], and likely contributes to normal glucose tolerance
Summary
Insulin-producing beta cells are the body’s central glucose sensors Key to their glucose sensing, is their dependence on lowaffinity glucose phosphorylation by glucokinase [1,2]. Intercellular variations in glucokinase abundance and activity give rise to proportionate variations in concentrationresponse curves for glucose-induced NAD(P)H, insulin synthesis and secretion [2,4,5,6,7]. This functional heterogeneity is subject to regulation in vivo – as shown in animals exposed to sulfonylurea [8]-, and likely contributes to normal glucose tolerance. Beta cell-selective proteins were identified through comparison with rat brain and liver tissue and with purified islet alpha cells, after geometrical normalization using 6 stably expressed reference proteins
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