The phorbol ester, 12-O-tetradecanoylphorbol 13-acetate (TPA), a potent stimulator of Erk, leads to the phosphorylation 4E-BP1 and its dissociation from eIF4E. In contrast agonists such as insulin, this occurs independently PKB activation. report, we investigate mechanism by which TPA regulates phosphorylation. Treatment HEK293 cells with was found result in at Ser64, Thr69, Thr36/45. TPA-stimulated all these sites is sensitive inhibitors MEK inhibitor mTOR, rapamycin, indicating that inputs...

125I protein labelling of oligo(dT)-selected RNA from feline calicivirus (FCV)-infected cells revealed that the genomic and 2.4 kb subgenomic RNAs FCV are linked to a 15 kDa (VPg). Proteinase K treatment RNA, remove VPg, led decrease in translatability but there was no obvious change site initiation. Addition cap analogue 7-methylGTP vitro translations had effect on translation suggesting is translated by cap-independent mechanism. Further evidence an unusual mechanism obtained translating...

Glucagon like peptide-1 (GLP1) is a Gs-coupled receptor agonist that exerts multiple effects on pancreatic β-cells, including the stimulation of insulin gene expression and secretion. In this report, we show treatment mouse β-cell line MIN6 with GLP1 leads to glucose-dependent activation Erk. These are mimicked by forskolin, direct activator adenylate cyclase, blocked H89, an inhibitor cAMP-dependent protein kinase. Additionally, provide evidence GLP1-stimulated Erk requires influx calcium...

Stimulation of serum-starved human embryonic kidney (HEK) 293 cells with either the phorbol ester, 12-<i>O</i>-tetradecanoylphorbol-13-acetate (TPA), or insulin resulted in increases phosphorylation 4E-BP1 and p70 S6 kinase, eIF4F assembly, protein synthesis. All these effects were blocked by rapamycin, a specific inhibitor mTOR. Phosphatidylinositol 3-kinase kinase B activated but not TPA. Therefore TPA can induce synthesis, independently both phosphatidylinositol B. Using two structurally...

Wasting of lean tissue as a consequence metabolic acidosis is serious problem in patients with chronic renal failure. A possible contributor inhibition by low pH the System (SNAT2) transporter, which carries amino acid L-glutamine (L-Gln) into muscle cells. The aim this study was to determine effect selective SNAT2 on intracellular profiles and acid-dependent signaling through mammalian target rapamycin L6 skeletal Inhibition competitive substrate methylaminoisobutyrate, (pH 7.1), or...

Rapamycin (sirolimus) is one of the primary immunosuppressants for islet transplantation. Yet there evidence that long-term treatment islet-transplant patients with rapamycin may be responsible subsequent loss graft function and viability. Therefore, objective this study was to elucidate molecular mechanism toxicity in beta cells.Experiments were performed on isolated rat human islets Langerhans MIN6 cells. The effects roles mammalian target complex 2 (mTORC2)/protein kinase B (PKB) cell...

cAMP and mTOR signalling pathways control a number of critical cellular processes including metabolism, protein synthesis, proliferation cell survival therefore understanding the events which integrate these two is particular interest. In this study, we show that pharmacological elevation [cAMP]i in mouse embryonic fibroblasts (MEFs) human kidney 293 (HEK293) cells inhibits mTORC1 activation via PKA-dependent mechanism. Although inhibitory effect on could be mediated by impinging cascades...

Age-related macular degeneration (AMD), a major cause of legal blindness in the elderly, is associated with genetic and environmental risk factors, such as cigarette smoking. Recent evidence shows that smoke (CS) contains high levels potent oxidants preferably targets retinal pigment epithelium (RPE) leading to oxidative damage apoptosis; however, mechanisms are poorly understood. The present study aimed investigate role endoplasmic reticulum (ER) stress unfolded protein response (UPR)...

Glucose regulates proinsulin biosynthesis via stimulation of the translation preproinsulin mRNA in pancreatic beta-cells. However, mechanism by which this occurs has remained unclear. Using recombinant adenoviruses that express with defined alterations, untranslated regions (UTRs) were examined for elements specifically control rat islets. These studies revealed 5'-UTR was necessary glucose translation, whereas 3'-UTR appeared to suppress translation. together 5'- and 3'-UTRs acted...

Feline calicivirus (FCV) is a small positive-stranded RNA virus within the family Caliciviridae. Its genome 7690 nucleotides in length and encodes three open reading frames (ORFs). The smallest, ORF3, located at extreme 3′ end of can potentially encode polypeptide approximately 12 kDa. In this paper, we report identification an ORF3-encoded FCV-infected cells using antiserum raised against bacterially-expressed bacteriophage T7 gene 10-ORF3 fusion protein. Although mRNA 0.5 kb, which could...

In the present study, we demonstrate that, in pancreatic β-cells, eIF2α (eukaryotic initiation factor 2α) phosphorylation response to a decrease glucose concentration is primarily mediated by activation of PERK [PKR (protein kinase RNA activated)-like endoplasmic reticulum kinase]. We provide evidence that this increase activity evoked energy status cell via potentially novel mechanism independent IRE1 (inositol requiring enzyme 1) and accumulation unfolded nascent proteins within reticulum....

Abstract Protein kinase R-like ER (PERK) is activated at physiologically low glucose concentrations in pancreatic β-cells. However, the molecular mechanisms by which PERK under these conditions and its role β-cell function are poorly understood. In this report, we investigated, dispersed rat islets of Langerhans mouse insulinoma-6 (MIN6) cells, relationship between extracellular concentration, free endoplasmic reticulum (ER) calcium concentration ([Ca2+]ER) measured directly using an...

The branched-chain amino acids (BCAA) leucine, isoleucine and valine, are essential that play a critical role in cellular signalling metabolism. They acutely stimulate insulin secretion activate the regulatory serine/threonine kinase mammalian target of rapamycin complex 1 (mTORC1), promotes increased β-cell mass function. effects BCAA on function dependent their active transport into cells via acid transporters thus expression activity these likely influence In this report, we show System-L...

Insulin resistance is a major cause of muscle wasting in patients with ESRD. Uremic metabolic acidosis impairs insulin signaling, which normally suppresses proteolysis. The low pH may inhibit the SNAT2 l-Glutamine (L-Gln) transporter, controls protein synthesis via amino acid-dependent signaling through mammalian target rapamycin (mTOR). Whether also regulates to pathways that control proteolysis unknown. In this study, inhibition selective competitive substrate methylaminoisobutyrate or (pH...

In pancreatic β-cells, glucose causes a rapid increase in the rate of protein synthesis. However, mechanism by which this occurs is poorly understood. report, we demonstrate, β-cell line MIN6, that stimulates recruitment ribosomes onto mRNA, indicative an initiation step This not mediated through availability complex eIF4F, because unable to stimulate eIF4F assembly or, absence amino acids, modulate phosphorylation status 4E-BP1. Moreover, MIN6 cells and isolated islets Langerhans,...

The accumulation of unfolded proteins within the endoplasmic reticulum (ER) causes ER stress and activation protein response (UPR). This can trigger ER-associated degradation autophagy, which clear restore homeostasis. Recently, it has become that ubiquitination plays an important role in regulation autophagy. In present study, we investigated how E3 ubiquitin ligase neural precursor cell-expressed, developmentally down-regulated 4-2 (Nedd4-2) interacts with mice, found increase expression...

In pancreatic β-cells, following an acute (within 1 h) increase in glucose concentration, there are rapid changes the expression of a large subset proteins. The change many these proteins is mediated by post-transcriptional mechanism through either increases or decreases rate translation from pre-existing transcripts. These proteins, whose synthesis rapidly up- down-regulated response to glucose, likely important mounting correct plasma concentrations. However, vast majority remain...

The objective of this study was to determine whether ER stress correlates with β-cell dysfunction in obesity-associated diabetes.Quantitative RT-PCR and western blot analysis were used investigate changes the expression markers stress, unfolded protein response (UPR) function islets isolated from (1) non-diabetic Zucker obese (ZO) female diabetic fatty (fZDF) rats compared their lean littermates (2) high-fat-diet-fed fZDF (HF-fZDF), induce diabetes, age-matched rats.Markers an adaptive...