ERp46 is reduced by high glucose and regulates insulin content in pancreatic β-cells

Abstract

Our studies focus on ERp46, an endoplasmic reticulum (ER) component, and analyze its involvement in glucose toxicity and in insulin production. Differences in pancreatic beta-TC-6 cell proteome under conditions of low vs. high glucose were examined by proteomic approaches, including two-dimensional gel electrophoresis, image analysis, and mass spectrometry. Among differentially expressed proteins, ERp46, a novel endoplasmic reticulum component, was examined further. The expression of ERp46 in pancreatic sections was analyzed by immunocytochemistry, and high glucose-induced alterations of expression were evaluated in cultured beta-cells, in isolated pancreatic islets, and in the pancreas of db/db diabetic animals. Inhibition of ERp46 expression by siRNA was performed to study its role in insulin production, in secretion, and in ER stress. Proteomic analysis led to identification of 46 differentially expressed spots corresponding to 23 proteins. Since ERp46 is a novel protein with a possible crucial role in secretory cells, we further analyzed its role in beta-cell function. ERp46 expression is reduced in high glucose concentration in beta-TC-6 cells and in isolated murine islets. Further analysis revealed high expression of ERp46 in pancreatic islets compared with exocrine tissue. Interestingly, a marked decrease in ERp46 expression was found in the pancreatic islets of db/db mice. Most importantly, siRNA-mediated knockdown of ERp46 in cultured beta-cells led to a significant decrease in the insulin content; however, no alterations in insulin mRNA levels were observed under these conditions. In addition, reduced expression of ERp46 by siRNA increased the expression of CHOP and peIF2a, indicating development of ER stress. We conclude that ERp46 may be an important component in the phenomenon of "glucose toxicity" involved in insulin production at the posttranslational level.

Proteomic profiling of HaCaT keratinocytes exposed to skin damaging detergents

“…Ряд белков, появившихся во всех опытных клетках, такие как TXNDC5, LRRC59 и SRSF2, ассоциированы с эндоплазматическим ретикулумом (ЭР) [24][25][26], что свидетельствует о его вовлечённости в реализацию ответа исследуемых клеток на токсическое повреждение. Известно, что одной из функций гладкого ЭР является обезвреживание различных токсических агентов [27].…”

Structure of the third catalytic domain of the protein disulfide isomerase ERp46

“…Previous studies have shown that human ERp46 acts as a stresssurvival factor (Sullivan et al, 2003) and as a regulator of insulin content in pancreatic cells (Alberti et al, 2009). It also specifically interacts with peroxiredoxin IV, a peroxidase in the ER (Jessop et al, 2009).…”

Papel da proteína dissulfeto isomerase na sinalização redox em células endoteliais e musculares lisas vasculares.

“…In all experiments, downregulation of Endo-PDI and PDI, however, had very similar effects although the siRNA used was demonstrated to act specifically. The uniform response to the downregulation of any of the two isomerases was very unexpected as the enzymes greatly differ in structure [6] and as individual substrate preferences have been demonstrated for proteins of the PDI family [14]. Also, differences with respect to their catalytic activity have been reported: Endo-PDI is a better thiol reductase than PDI, whereas PDI has higher isomerase activity [19].…”

“…It can therefore only be speculated why functionally the two proteins were so equally important. Despite the considerations mentioned above, the substrates of PDI and Endo-PDI overlap significantly [14]. It may therefore be that some reductase or isomerase threshold activity has to be reached to maintain activity of the pathway.…”

“…Owing to the generation of autacoids the vascular endothelium also falls into this category and thus expresses high levels of PDI and several PDI homologs. One of these is Endo-PDI [13], also termed ERp46 [8], plasma cell thioredoxin-related protein, or thioredoxin domaincontaining protein 5 precursor, which is expressed in several cells such as pancreatic β cells [14] and plasma cells [15], but is most prominent in the vascular endothelium [13]. Because of the high endothelial expression, the protein was first discovered in these cells.…”