Increased hexosamine biosynthetic pathway flux dedifferentiates INS-1E cells and murine islets by an extracellular signal-regulated kinase (ERK)1/2-mediated signal transmission pathway.

AIMS/HYPOTHESIS: Beta cell failure is caused by loss of cell mass, mostly by apoptosis, but also by simple dysfunction (decline of glucose-stimulated insulin secretion, downregulation of specific gene expression). Apoptosis and dysfunction are caused, at least in part, by lipoglucotoxicity. The mechanisms implicated are oxidative stress, increase in the hexosamine biosynthetic pathway (HBP) flux and endoplasmic reticulum (ER) stress. Oxidative stress plays a role in glucotoxicity-induced beta cell dedifferentiation, while glucotoxicity-induced ER stress has been mostly linked to beta cell apoptosis. We sought to clarify whether ER stress caused by increased HBP flux participates in a dedifferentiating response of beta cells, in the absence of relevant apoptosis. METHODS: We used INS-1E cells and murine islets. We analysed the unfolded protein response and the expression profile of beta cells by real-time RT-PCR and western blot. The signal transmission pathway elicited by ER stress was investigated by real-time RT-PCR and immunofluorescence. RESULTS: Glucosamine and high glucose induced ER stress, but did not decrease cell viability in INS-1E cells. ER stress caused dedifferentiation of beta cells, as shown by downregulation of beta cell markers and of the transcription factor, pancreatic and duodenal homeobox 1. Glucose-stimulated insulin secretion was inhibited. These effects were prevented by the chemical chaperone, 4-phenyl butyric acid. The extracellular signal-regulated kinase (ERK) signal transmission pathway was implicated, since its inhibition prevented the effects induced by glucosamine and high glucose. CONCLUSIONS/INTERPRETATION: Glucotoxic ER stress dedifferentiates beta cells, in the absence of apoptosis, through a transcriptional response. These effects are mediated by the activation of ERK1/2.

Cell fate following ER stress: just a matter of "quo ante" recovery or death?

The endoplasmic reticulum (ER) is a complex and multifunctional organelle. It is the intracellular compartment of protein folding, a complex task, both facilitated and monitored by ER folding enzymes and molecular chaperones. The ER is also a stress-sensing organelle. It senses stress caused by disequilibrium between ER load and folding capacity and responds by activating signal transduction pathways, known as unfolded protein response (UPR). Three major classes of transducer are known, inositol-requiring protein-1 (IRE1), activating transcription factor-6 (ATF6), and protein kinase RNA (PKR)-like endoplasmic reticulum kinase (PERK), which sense with their endoluminal domain the state of protein folding, although the exact mechanism(s) involved is not entirely clear. Depending on whether the homeostatic response of the UPR is successful in restoring an equilibrium between ER load and protein folding or not, the two possible outcomes of the UPR so far considered have been life or death. Indeed, recent efforts have been devoted to understand the life/death switch mechanisms. However, recent data suggest that what appears to be a pure binary decision may in fact be more complex, and survival may be achieved at the expenses of luxury cell functions, such as expression of differentiation genes.

Two sunflower 17.6HSP genes, arranged in tandem and highly homologous, are induced differently by various elicitors.

Plants respond to environmental stimuli, such as heat shock, by re-programming cellular activity through differential gene expression, mainly controlled at the transcription level. The current study refers to two sunflower small heat shock protein (sHSP) genes arranged in tandem in head-to-head orientation and linked by a 3809 bp region. These genes exhibit only slight structural differences in the coding portion. They code for cytosolic class I sHSPs and are named HaHSP17.6a and HaHSP17.6b according to the molecular weight of the putative proteins. The genomic organization of these genes is consistent with the idea that many HSP genes originate from duplication events; in this case, probably an inversion and duplication occurred. The HaHSP17.6a and HaHSP17.6b genes are characterized by different expression levels under various heat stress conditions; moreover, their expression is differently induced by various elicitors. The differential regulation observed for HaHSP17.6a and HaHSP17.6b genes differs from previous observations on duplicated sHSP genes in plants.

Isolation and characterisation of a cDNA for a novel small HSP from sunflower suspension cell cultures.

A full-length cDNA encoding a novel small heat shock protein (HaHSP17.9) was isolated from a cDNA library of sunflower (Helianthus annuus cv. Gloriasol). The deduced amino acid sequence exhibited high degree homology to the class I cytosolic sHSPs from other plant species, and contained all the conserved regions characteristic of this class of proteins. Northern analyses showed that the transcript homologous to HaHSP17.9 accumulates during heat shock in suspension cultured cells and in the different parts of sunflower seedlings.