Deletion of SERP1/RAMP4, a component of the endoplasmic reticulum (ER) translocation sites, leads to ER stress.

Stress-associated endoplasmic reticulum (ER) protein 1 (SERP1), also known as ribosome-associated membrane protein 4 (RAMP4), is a Sec61-associated polypeptide that is induced by ER stress. SERP1-/- mice, made by targeted gene disruption, demonstrated growth retardation, increased mortality, and impaired glucose tolerance. Consistent with high levels of SERP1 expression in pancreas, pancreatic islets from SERP1-/- mice failed to rapidly synthesize proinsulin in response to a glucose load. In addition, reduced size and enhanced ER stress were observed in the anterior pituitary of SERP1-/- mice, and growth hormone production was slowed in SERP1-/- pituitary after insulin stimulation. Experiments using pancreatic microsomes revealed aberrant association of ribosomes and the Sec61 complex and enhanced ER stress in SERP1-/- pancreas. In basal conditions, the Sec61 complex in SERP1-/- microsomes was more cofractionated with ribosomes, compared with SERP1+/+ counterparts, in high-salt conditions. In contrast, after glucose stimulation, the complex showed less cofractionation at an early phase (45 min) but more at a later phase (120 min). Although intracellular insulin/proinsulin levels were not significantly changed in both genotypes, these results suggest that subtle changes in translocation efficiency play an important role in the regulation of ER stress and rapid polypeptide synthesis.

The endoplasmic reticulum chaperone improves insulin resistance in type 2 diabetes.

To determine the role of the endoplasmic reticulum (ER) in diabetes, Akita mice, a mouse model of type 2 diabetes, were mated with either heterozygous knockout mice or two types of transgenic mice of 150-kDa oxygen-regulated protein (ORP150), a molecular chaperone located in the ER. Systemic expression of ORP150 in Akita mice improves insulin intolerance, whereas the exclusive overexpression of ORP150 in pancreatic beta-cells of Akita mice did not change their glucose tolerance. Both an insulin tolerance test and hyperinsulinemic-euglycemic clamp revealed that ORP150 enhanced glucose uptake, accompanied by suppression of oxidized protein. Furthermore, ORP150 enhanced the insulin sensitivity of myoblast cells treated with hydrogen peroxide. These data suggest that ORP150 plays an important role in insulin sensitivity and is a potential target for the treatment of diabetes.

Expression of 150-kd oxygen-regulated protein in the hippocampus suppresses delayed neuronal cell death.

ORP150-150-kd oxygen-regulated protein-is a novel stress protein localized in the endoplasmic reticulum (ER). To investigate the role of ORP150 in delayed neuronal cell death, the authors examined its expression in the gerbil brain after an ischemic insult. The expression of ORP150 antigen, as well as its transcripts, was observed in the CA1 region after the occlusion of the common carotid artery, and the preconditioning enhanced this expression. In cultured neurons, exposure either to hypoxia or to glutamate induced the expression of ORP150, and this effect was also observed by treating the culture with breferdin A or thapsigargin, indicating that both glutamate and hypoxia can cause stress in the ER (ER stress). Neurons became more vulnerable to these stresses following treatment with cycloheximide or after infection with an adenovirus carrying the ORP150-antisense structure. In contrast, the overexpression of ORP150 by an adenovirus suppressed neuronal cell death, and this was accompanied by the suppression of Ca2+ elevation and proteolytic activity induced by glutamate. Further, overexpression of ORP150 in CA1 neurons by an adenovirus carrying the ORP150-sense structure suppressed delayed neuronal cell death after ischemia. These data suggest a possible function of ORP150 as an intracellular apparatus that participates in a protective response in ischemic tolerance.

Transmission of cell stress from endoplasmic reticulum to mitochondria: enhanced expression of Lon protease.

The rat homologue of a mitochondrial ATP-dependent protease Lon was cloned from cultured astrocytes exposed to hypoxia. Expression of Lon was enhanced in vitro by hypoxia or ER stress, and in vivo by brain ischemia. These observations suggested that changes in nuclear gene expression (Lon) triggered by ER stress had the potential to impact important mitochondrial processes such as assembly and/or degradation of cytochrome c oxidase (COX). In fact, steady-state levels of nuclear-encoded COX IV and V were reduced, and mitochondrial-encoded subunit II was rapidly degraded under ER stress. Treatment of cells with cycloheximide caused a similar imbalance in the accumulation of COX subunits, and enhanced mRNA for Lon and Yme1, the latter another mitochondrial ATP-dependent protease. Furthermore, induction of Lon or GRP75/mtHSP70 by ER stress was inhibited in PERK (-/-) cells. Transfection studies revealed that overexpression of wild-type or proteolytically inactive Lon promoted assembly of COX II into a COX I-containing complex, and partially prevented mitochondrial dysfunction caused by brefeldin A or hypoxia. These observations demonstrated that suppression of protein synthesis due to ER stress has a complex effect on the synthesis of mitochondrial-associated proteins, both COX subunits and ATP-dependent proteases and/or chaperones contributing to assembly of the COX complex.