Endoplasmic Reticulum Stress: Its Role in Disease and Novel Prospects for Therapy

Abstract

The endoplasmic reticulum (ER) is a multifunctional organelle required for lipid biosynthesis, calcium storage, and protein folding and processing. A number of physiological and pathological conditions, as well as a variety of pharmacological agents, are able to disturb proper ER function and thereby cause ER stress, which severely impairs protein folding and therefore poses the risk of proteotoxicity. Specific triggers for ER stress include, for example, particular intracellular alterations (e.g., calcium or redox imbalances), certain microenvironmental conditions (e.g., hypoglycemia, hypoxia, and acidosis), high-fat and high-sugar diet, a variety of natural compounds (e.g., thapsigargin, tunicamycin, and geldanamycin), and several prescription drugs (e.g., bortezomib/Velcade, celecoxib/Celebrex, and nelfinavir/Viracept). The cell reacts to ER stress by initiating a defensive process, called the unfolded protein response (UPR), which is comprised of cellular mechanisms aimed at adaptation and safeguarding cellular survival or, in cases of excessively severe stress, at initiation of apoptosis and elimination of the faulty cell. In recent years, this dichotomic stress response system has been linked to several human diseases, and efforts are underway to develop approaches to exploit ER stress mechanisms for therapy. For example, obesity and type 2 diabetes have been linked to ER stress-induced failure of insulin-producing pancreatic beta cells, and current research efforts are aimed at developing drugs that ameliorate cellular stress and thereby protect beta cell function. Other studies seek to pharmacologically aggravate chronic ER stress in cancer cells in order to enhance apoptosis and achieve tumor cell death. In the following, these principles will be presented and discussed.

Altered Activation of Protein Kinase PKR and Enhanced Apoptosis in Dystonia Cells Carrying a Mutation in PKR Activator Protein PACT

“…dsRNA Binding Assay-The in vitro translated, 35 S-labeled PACT proteins were synthesized using the TNT-T7-coupled reticulocyte lysate system from Promega, and the dsRNA binding activity was measured by using the previously established poly(I)⅐poly(C)-agarose binding assay (3,11). 4 l of in vitro translation products were diluted with 25 l of binding buffer (20 mM Tris, pH 7.5, 0.3 M NaCl, 5 mM MgCl 2 , 1 mM DTT, 0.1 mM PMSF, 0.5% Nonidet P-40, 10% glycerol) and incubated with 25 l of poly(I)⅐poly(C)-agarose beads at 30°C for 30 min.…”

“…For a competition assay with soluble single-or double-stranded RNA, 1 g of poly(C) or poly(I)⅐poly(C) was incubated with the proteins for 15 min at 30°C before the addition of poly(I)⅐poly(C)-agarose beads. To ascertain specific interactions between PACT proteins and poly(I)⅐poly(C)-agarose beads, in vitro translated, 35 S-labeled firefly luciferase protein was assayed for binding to the poly(I)⅐poly(C)-agarose beads using same conditions. The T lanes represent total radioactive proteins in the reticulocyte lysate, and the B lanes represent the proteins that remain bound to poly(I)⅐poly(C)-agarose beads after washing.…”

“…Co-immunoprecipitation with in Vitro Translated ProteinsIn vitro translated, 35 S-labeled PKR, and FLAG epitope-tagged PACT proteins were synthesized using the TNT-T7 coupled reticulocyte system from Promega, and the co-immunoprecipitation assay was performed as described before (11).…”

Role of endoplasmic reticulum stress in drug‐induced toxicity

“…More precisely, thapsigargin induces ER stress by inhibiting sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) (Fig. 2), thus leading to severe depletion of ER calcium (Kaufman 1999; Denmeade and Isaacs 2005; Schönthal 2012). In addition to ER stress‐induced activation of downstream effectors that can trigger cell death, the concomitant increase in free cytosolic calcium is also a potent proapoptotic signal in different types of cells (Jiang et al.…”

“…Notably, this biosynthetic step takes place within the ER in eukaryotic cells (Bieberich 2014). Consequently, impairment of glycosylation of newly synthesized proteins is leading to the disruption of their folding in the ER (Kaufman 1999; Schönthal 2012). In addition, this alteration of peptide conformation can secondarily prevent the cellular secretion of some proteins such as different immunoglobulins (Hickman et al.…”