Developing Vanadium as an Antidiabetic or Anticancer Drug: A Clinical and Historical Perspective.

Vanadium has been known for centuries to have beneficial effects on health and has the potential to be used as an alternative to other diabetic and anticancer medicines. The beneficial effects of vanadium salts or organic compounds have been explored in vitro, ex vivo, and in vivo in animal and human studies. A consensus among researchers is that increased bioavailability of these compounds could markedly increase the efficacy of this class of compounds.

Knockout of USP19 Deubiquitinating Enzyme Prevents Muscle Wasting by Modulating Insulin and Glucocorticoid Signaling.

Muscle atrophy arises because of many chronic illnesses, as well as from prolonged glucocorticoid treatment and nutrient deprivation. We previously demonstrated that the USP19 deubiquitinating enzyme plays an important role in chronic glucocorticoid- and denervation-induced muscle wasting. However, the mechanisms by which USP19 exerts its effects remain unknown.

Insulin Signalling: The Inside Story.

Insulin signalling begins with binding to its cell surface insulin receptor (IR), which is a tyrosine kinase. The insulin receptor kinase (IRK) is subsequently autophosphorylated and activated to tyrosine phosphorylate key cellular substrates that are essential for entraining the insulin response. Although IRK activation begins at the cell surface, it is maintained and augmented following internalization into the endosomal system (ENS).

Spatial and Temporal Regulation of Receptor Tyrosine Kinase Activation and Intracellular Signal Transduction.

Epidermal growth factor (EGF) and insulin receptor tyrosine kinases (RTKs) exemplify how receptor location is coupled to signal transduction. Extracellular binding of ligands to these RTKs triggers their concentration into vesicles that bud off from the cell surface to generate intracellular signaling endosomes. On the exposed cytosolic surface of these endosomes, RTK autophosphorylation selects the downstream signaling proteins and lipids to effect growth factor and polypeptide hormone action.

Assessment of internalization and endosomal signaling: studies with insulin and EGF.

Endosomes are isolated from rat liver using high-speed centrifugation through sucrose density gradients. They are distinguishable from Golgi elements, with which they coisolate, by their capacity to concentrate internalized protein ligands (viz., insulin and epidermal growth factor (EGF)) in receptor-bound intact form.

Epidermal growth factor-induced vacuolar (H+)-atpase assembly: a role in signaling via mTORC1 activation.

Using proteomics and immunofluorescence, we demonstrated epidermal growth factor (EGF) induced recruitment of extrinsic V(1) subunits of the vacuolar (H(+))-ATPase (V-ATPase) to rat liver endosomes. This was accompanied by reduced vacuolar pH. Bafilomycin, an inhibitor of V-ATPase, inhibited EGF-stimulated DNA synthesis and mammalian target of rapamycin complex 1 (mTORC1) activation as indicated by a decrease in eukaryotic initiation factor 4E-binding 1 (4E-BP1) phosphorylation and p70 ribosomal S6 protein kinase (p70S6K) phosphorylation and kinase activity.

Global analysis of protein phosphorylation networks in insulin signaling by sequential enrichment of phosphoproteins and phosphopeptides.

Although the important role of protein phosphorylation in insulin signaling networks is well recognized, its analysis in vivo has not been pursued in a systematic fashion through proteome-wide studies. Here we undertake a global analysis of insulin-induced changes in the rat liver cytoplasmic and endosomal phosphoproteome by sequential enrichment of phosphoproteins and phosphopeptides. After subcellular fractionation proteins were denatured and loaded onto iminodiacetic acid-modified Sepharose with immobilized Al³⁺ ions (IMAC-Al resin).

Cellular signalling: Peptide hormones and growth factors.

Peptide hormones and growth factors initiate signalling by binding to and activating their cell surface receptors. The activated receptors interact with and modulate the activity of cell surface enzymes and adaptor proteins which entrain a series of reactions leading to metabolic and proliferative signals. Rapid internalization of ligand-receptor complexes into the endosomal system both prolongs and augments events initiated at the cell surface.

Compartmentalization of EGFR in cellular membranes: role of membrane rafts.

There is now abundant evidence that the intracellular concentration of the EGFR and many other receptors for peptide hormones and growth factors is important for the temporal and spatial regulation of cell signaling. Spatial control is achieved by the selective compartmentalization of signaling components into endosomes. However further control may be effected by sequestration into sub-domains within a given organelle such as membrane rafts which are dynamic, nano scale structures rich in cholesterol and sphingolipids.

Compartmentalization of epidermal growth factor receptor in liver plasma membrane.

We have investigated epidermal growth factor (EGF)-induced compartmentalization and activation of the EGF receptor (EGFR) in rat liver plasma membrane (PM) raft subfractions prepared by three different biochemical methods previously developed to characterize the composition of membrane rafts. Only detergent-resistant membranes (DRMs) possessed the basic characteristics attributed to membrane rafts. Following the administration of a low dose of EGF (1 microg/100 g BW) the content of EGFR in PM-DRMs did not change significantly; whereas after a higher dose of EGF (5 microg/100 g BW) we observed a rapid and marked disappearance of EGFR (around 80%) from both PM and DRM fractions.

Quantitative analysis of a proteome by N-terminal stable-isotope labelling of tryptic peptides.

Covalent modification of peptides and proteins with compounds containing stable isotopes (isotope tagging) has become an essential tool to detect dynamic changes in the proteome following external or internal influence; however, using terminal amino groups for global isotope labelling of tryptic peptides is challenged by the similar reactivity of the amino groups of lysine residues. We describe a new quantitative method based on selective tagging of the terminal amino groups of tryptic peptides with pentafluorophenyl esters containing stable isotopes. The labelled peptides were resolved by two-dimensional nanoflow liquid chromatography on weak anion-exchange and reversed-phase columns and then identified and quantified by tandem mass spectrometry.

Compartmentalization of signaling-competent epidermal growth factor receptors in endosomes.

In this study, the preparation of detergent-resistant membranes (DRMs) and the immunoisolation of intracellular vesicles enriched in raft markers were used to investigate the effect of physiological doses of epidermal growth factor (EGF) in vivo on the compartmentalization and activation of EGF receptor (EGFR) in rat liver endosomes. Both of these techniques show that after EGF administration, a distinctive population of intracellular EGFR, which was characterized by a high level of tyrosine phosphorylation, accumulated in endosomes. EGFR recruited to early endosomes were more tyrosine phosphorylated than those from late endosomes.

A genome-wide association study identifies novel risk loci for type 2 diabetes.

Type 2 diabetes mellitus results from the interaction of environmental factors with a combination of genetic variants, most of which were hitherto unknown. A systematic search for these variants was recently made possible by the development of high-density arrays that permit the genotyping of hundreds of thousands of polymorphisms. We tested 392,935 single-nucleotide polymorphisms in a French case-control cohort.

Transcriptional regulation by insulin: from the receptor to the gene.

Insulin, after binding to its receptor, regulates many cellular processes and the expression of several genes. For a subset of genes, insulin exerts a negative effect on transcription; for others, the effect is positive. Insulin controls gene transcription by modifying the binding of transcription factors on insulin-response elements or by regulating their transcriptional activities.

Regulation of hepatic insulin-like growth factor-binding protein-1 gene expression by insulin: central role for mammalian target of rapamycin independent of forkhead box O proteins.

The expression of IGF-binding protein-1 (IGFBP-1) is induced in rat liver by dexamethasone and glucagon and is completely inhibited by 100 nM insulin. Various studies have implicated phosphatidylinositol 3-kinase, protein kinase B (Akt), phosphorylation of the transcription factors forkhead in rhabdomyosarcoma 1 (Foxo1)/Foxo3, and the mammalian target of rapamycin (mTOR) in insulin's effect. In this study we examined insulin regulation of IGFBP-1 in both subconfluent and confluent hepatocytes.

Insulin receptor kinase-associated phosphotyrosine phosphatases in hepatic endosomes: assessing the role of phosphotyrosine phosphatase-1B.

Previous work has shown that bisperoxo(1,10-phenanthroline)-oxovanadate(v) anion [bpV(phen)] induces potent insulin-mimicking effects in the rat, selectively activates the endosomal (EN) insulin receptor kinase (IRK) in liver, and markedly abolishes endosomal IRK-associated phosphotyrosine phosphatase (PTP) activity while reducing that of total ENs by approximately 30%. In this study we examined the relatively selective effect of bpv(phen) on endosomal PTP activities for the purpose of defining IRK-associated PTP(s). Using an in-gel PTP assay, we detected multiple (approximately 20) species of endosomal PTP (30 to >220 kDa), with five that were markedly inhibited after in vivo bpV(phen) administration.

The G protein-coupled receptor kinase-2 is a TGFbeta-inducible antagonist of TGFbeta signal transduction.

Signaling from the activin/transforming growth factor beta (TGFbeta) family of cytokines is a tightly regulated process. Disregulation of TGFbeta signaling is often the underlying basis for various cancers, tumor metastasis, inflammatory and autoimmune diseases. In this study, we identify the protein G-coupled receptor kinase 2 (GRK2), a kinase involved in the desensitization of G protein-coupled receptors (GPCR), as a downstream target and regulator of the TGFbeta-signaling cascade.

Effect of insulin on caveolin-enriched membrane domains in rat liver.

Compartmentalization of signaling molecules may explain, at least in part, how insulin or growth factors achieve specificity. Caveolae/rafts are specialized lipid compartments that have been implicated in insulin signaling. In the present study, we investigated the role of caveolin-enriched membrane domains (CMD) in mediating insulin signaling in rat liver.

Effect of inhibiting vacuolar acidification on insulin signaling in hepatocytes.

Previous studies have shown that the endosomal apparatus plays an important role in insulin signaling. Inhibition of endosomal acidification leads to a decrease in insulin-insulin receptor kinase (IRK) dissociation and insulin degradation. Thus, vacuolar pH could function as a modulator of insulin signaling in endosomes.

Regulation of insulin receptor kinase activity by endosomal processes: possible areas for therapeutic intervention.

The insulin receptor kinase (IRK) is activated following insulin binding and is rapidly internalized into endosomes (ENs) from which signaling occurs. Four endosomal processes limit the intensity and duration of intracellular signal transduction: (i) insulin degradation by an endosomal acidic insulinase, cathepsin D, which removes the ligand leading to receptor deactivation; (ii) IRK dephosphorylation by an associated protein tyrosine phosphatase abrogates its activated state; (iii) acidification of ENs changes IRK conformation reducing its affinity for ligand and inactivating its kinase; and (iv) trafficking within ENs can sequester activated IRK from signal transduction elements. Each process presents an opportunity for new potential therapeutic approaches.

Gab2 tyrosine phosphorylation by a pleckstrin homology domain-independent mechanism: role in epidermal growth factor-induced mitogenesis.

In primary rat hepatocyte cultures, activation of phosphatidylinositol 3-kinase is both necessary and sufficient to account for epidermal growth factor (EGF)-induced DNA synthesis. In these cells, three major p85-containing complexes were formed after EGF treatment: ErbB3-p85, Shc-p85, and a multimeric Gab2-Grb2-SHP2-p85, which accounted for more than 80% of total EGF-induced PI3K activity (Kong, M., C.

Different Forms of Vanadate on Sugar Transport in Insulin Target and Nontarget Cells.

The effects of several vanadates (ie, orthovanadate, pervanadate, and two stable peroxovanadium compounds) on basal and insulin-stimulated 2-DG transport in insulin target and nontarget cell lines are reported, herein. In nontarget cells, exposure to vanadates (5 x 10(-6) to 10(-4) mol/L) resulted in 2-DG transport stimulatory responses similar to those observed in 2-DG transport post exposure to 667 nmol/L insulin alone, or insulin in combination with vanadates. In 3T3-L1 adipocytes and L6 myotubes, exposure to a vanadate compound or 67 nmol/L insulin, stimulated 2-DG transport dramatically.

Epidermal growth factor-induced DNA synthesis. Key role for Src phosphorylation of the docking protein Gab2.

We have previously demonstrated that phosphatidylinositol 3-kinase (PI3-kinase) is necessary and sufficient to account for epidermal growth factor (EGF)-induced mitogenesis in rat primary hepatocytes. A cytosolic Gab2-containing complex accounts for >80% of the total EGF-induced PI3-kinase activity (Kong, M., Mounier, C.