Adaptive preconditioning in neurological diseases - therapeutic insights from proteostatic perturbations.

In neurological disorders, both acute and chronic neural stress can disrupt cellular proteostasis, resulting in the generation of pathological protein. However in most cases, neurons adapt to these proteostatic perturbations by activating a range of cellular protective and repair responses, thus maintaining cell function. These interconnected adaptive mechanisms comprise a 'proteostasis network' and include the unfolded protein response, the ubiquitin proteasome system and autophagy. Interestingly, several recent studies have shown that these adaptive responses can be stimulated by preconditioning treatments, which confer resistance to a subsequent toxic challenge - the phenomenon known as hormesis. In this review we discuss the impact of adaptive stress responses stimulated in diverse human neuropathologies including Parkinson׳s disease, Wolfram syndrome, brain ischemia, and brain cancer. Further, we examine how these responses and the molecular pathways they recruit might be exploited for therapeutic gain. This article is part of a Special Issue entitled SI:ER stress.

Activation of OASIS family, ER stress transducers, is dependent on its stabilization.

Endoplasmic reticulum (ER) stress transducers transduce signals from the ER to the cytoplasm and nucleus when unfolded proteins accumulate in the ER. BBF2 human homolog on chromosome 7 (BBF2H7) and old astrocyte specifically induced substance (OASIS), ER-resident transmembrane proteins, have recently been identified as novel ER stress transducers that have roles in chondrogenesis and osteogenesis, respectively. However, the molecular mechanisms that regulate the activation of BBF2H7 and OASIS under ER stress conditions remain unresolved. Here, we showed that BBF2H7 and OASIS are notably unstable proteins that are easily degraded via the ubiquitin-proteasome pathway under normal conditions. ER stress conditions enhanced the stability of BBF2H7 and OASIS, and promoted transcription of their target genes. HMG-CoA reductase degradation 1 (HRD1), an ER-resident E3 ubiquitin ligase, ubiquitinated BBF2H7 and OASIS under normal conditions, whereas ER stress conditions dissociated the interaction between HRD1 and BBF2H7 or OASIS. The stabilization of OASIS in Hrd1(-/-) cells enhanced the expression of collagen fibers during osteoblast differentiation, whereas a knockdown of OASIS in Hrd1(-/-) cells suppressed the production of collagen fibers. These findings suggest that ER stress stabilizes OASIS family members and this is a novel molecular mechanism for the activation of ER stress transducers.

Mutations in the core and NS5A region of hepatitis C virus genotype 1b and correlation with response to pegylated-interferon-alpha 2b and ribavirin combination therapy.

Mutations in two regions of hepatitis C virus (HCV) have been implicated in influencing response to interferon (IFN) therapy. Substitutions in the NS5A region of HCV have been associated with response to IFN therapy, and this region has been known as the IFN sensitivity-determining region (ISDR). The mutations in the core region of HCV have also been reported to predict IFN response. The aim of this study was to investigate whether amino acid substitutions in the core region and ISDR among patients with HCV genotype 1b affect the response to IFN therapy. A total of 213 patients who completed IFN treatment were randomly selected. All patients received pegylated-IFN-alpha 2b once each week, plus oral ribavirin daily for 48 weeks. Of the 213 patients, 117 (54.9%) showed early virologic response (EVR), with HCV-negativity, at 12 weeks. Factors related to EVR on multivariate analysis were non-Gln70 and Leu91 in the core region, and ISDR mutant-type. One hundred and two (47.9%) showed a sustained virologic response (SVR). SVR occurred more frequently in patients without Gln70 (55.4%) than in those with Gln70 (21.3%) (P < 0.0001). SVR was achieved in 43.6% of patients with wild-type ISDR and 62.5% of patients with mutant-type (P = 0.0227). Of the 34 patients who simultaneously had non-Gln70 and mutant-type ISDR, 26 (76.5%) achieved SVR. Factors related to SVR on multivariate analysis were non-Gln70 and ISDR mutant-type. In conclusion, amino acid substitutions in the core region and ISDR were useful for predicting the response to IFN in patients with HCV genotype 1b.

A switch from life to death in endoplasmic reticulum stressed ß-cells.

ß-Cell death is an important pathogenic component of both type 1 and type 2 diabetes. Recent findings indicate that cell signalling pathways emanating from the endoplasmic reticulum (ER) play an important role in the regulation of ß-cell death during the progression of diabetes. Homeostasis within the ER must be maintained to produce properly folded secretory proteins, such as insulin, in response to the body's need for them. However, the sensitive protein-folding environment in the ER can be perturbed by genetic and environmental factors leading to ER stress. To counteract ER stress, ß-cells activate cell signalling pathways termed the unfolded protein response (UPR). The UPR functions as a binary switch between life and death, regulating both survival and death effectors. The outcome of this switch depends on the nature of the ER stress condition, the regulation of UPR activation and the expression and activation of survival and death components. This review discusses the mechanisms and the components in this switch and highlights the roles of this UPR's balancing act between life and death in ß-cells.

AATF mediates an antiapoptotic effect of the unfolded protein response through transcriptional regulation of AKT1.

Endoplasmic reticulum (ER) stress-mediated cell death has an important role in the pathogenesis of chronic diseases, including diabetes and neurodegeneration. Although proapoptotic programs activated by ER stress have been extensively studied, identification and characterization of antiapoptotic programs that counteract ER stress are currently incomplete. Through the gene expression profiling of beta-cells lacking Wolfram syndrome 1 gene (WFS1), a causative gene for Wolfram syndrome, we discovered a novel antiapoptotic gene of the unfolded protein response (UPR), apoptosis antagonizing transcription factor (AATF). Here, we study the regulation of AATF, identify its target genes, and determine the basis for its antiapoptotic activities in response to ER stress. We show that AATF is induced by ER stress through the PERK-eIF2alpha pathway and transcriptionally activates the v-akt murine thymoma viral oncogene homolog 1 (AKT1) gene through signal transducer and activator of transcription 3 (Stat3), which sustains Akt1 activation and promotes cell survival. Ectopic expression of AATF or a constitutively active form of AKT1 confers on cells resistance to ER stress-mediated cell death, whereas RNAi-mediated knockdown of AATF or AKT1 renders cells sensitive to ER stress. We also discovered a positive crosstalk between the AATF and WFS1 signaling pathways. Thus, WFS1 deficiency or AATF deficiency mediates a self-perpetuating cycle of cell death. Our results reveal a novel antiapoptotic program relevant to the treatment of diseases caused by ER stress-mediated cell death.

Amyloid beta induces neuronal cell death through ROS-mediated ASK1 activation.

Amyloid beta (Abeta) is a main component of senile plaques in Alzheimer's disease and induces neuronal cell death. Reactive oxygen species (ROS), nitric oxide and endoplasmic reticulum (ER) stress have been implicated in Abeta-induced neurotoxicity. We have reported that apoptosis signal-regulating kinase 1 (ASK1) is required for ROS- and ER stress-induced JNK activation and apoptosis. Here we show the involvement of ASK1 in Abeta-induced neuronal cell death. Abeta activated ASK1 mainly through production of ROS but not through ER stress in cultured neuronal cells. Importantly, ASK1-/- neurons were defective in Abeta-induced JNK activation and cell death. These results indicate that ROS-mediated ASK1 activation is a key mechanism for Abeta-induced neurotoxicity, which plays a central role in Alzheimer's disease.

Neuroleptic malignant syndrome with prolonged catatonia in a dopa-responsive dystonia patient.

The authors describe a patient with dopa-responsive dystonia who developed neuroleptic malignant syndrome with prolonged catatonia following treatment with neuroleptic agents. Use of these agents probably expanded the patient's neuronal dysfunction beyond the nigrostriatal system to involve multiple dopaminergic systems. Electroconvulsive treatment alleviated the prolonged catatonia.

Delayed delivery of AAV-GDNF prevents nigral neurodegeneration and promotes functional recovery in a rat model of Parkinson's disease.

Glial cell line-derived neurotrophic factor (GDNF) is a strong candidate agent in the neuroprotective treatment of Parkinson's disease (PD). We investigated whether adeno-associated viral (AAV) vector-mediated delivery of a GDNF gene in a delayed manner could prevent progressive degeneration of dopaminergic (DA) neurons, while preserving a functional nigrostriatal pathway. Four weeks after a unilateral intrastriatal injection of 6-hydroxydopamine (6-OHDA), rats received injection of AAV vectors expressing GDNF tagged with FLAG peptide (AAV-GDNFflag) or beta-galactosidase (AAV-LacZ) into the lesioned striatum. Immunostaining for FLAG demonstrated retrograde transport of GDNFflag to the substantia nigra (SN). The density of tyrosine hydroxylase (TH)-positive DA fibers in the striatum and the number of TH-positive or cholera toxin subunit B (CTB, neuronal tracer)-labeled neurons in the SN were significantly greater in the AAV-GDNFflag group than in the AAV-LacZ group. Dopamine levels and those of its metabolites in the striatum were remarkably higher in the AAV-GDNFflag group compared with the control group. Consistent with anatomical and biochemical changes, significant behavioral recovery was observed from 4-20 weeks following AAV-GDNFflag injection. These data indicate that a delayed delivery of GDNF gene using AAV vector is efficacious even 4 weeks after the onset of progressive degeneration in a rat model of PD.

Gastrin and Helicobacter pylori in low-grade MALT lymphoma patients.

BACKGROUND: This study of patients with Helicobacter pylori infection and low-grade MALT lymphoma aimed to investigate: 1) the effect of H. pylori eradication therapy on the serum gastrin level, 2) whether changes of the serum gastrin level after therapy could predict the prognosis of patients with this tumour, and 3) the relationship between the gastric H. pylori load, the serum gastrin level and the status of MALT lymphoma. METHODS: Thirteen patients with documented low-grade MALT lymphoma and H. pylori infection were enrolled and received H. pylori eradication therapy as the sole initial treatment. The presence of H. pylori, the serum gastrin level, the endoscopic findings, the pathologic features of the biopsies and resected specimens, and the endoscopic ultrasonography findings were evaluated before and after therapy. Follow-up was carried out every 3-6 months. RESULTS: H. pylori eradication was eventually achieved in all 13 patients. The pretreatment fasting serum gastrin level decreased from 177.1 +/- 107.4 pg/ml to 129.2 +/- 78.1, 96.4 +/- 66.6 and 80.1 +/- 42.7 pg/ml after 0-3, 3-6 and 6-9 months, respectively (all P < 0.05). Successful eradication of H. pylori was followed by a decrease of the fasting serum gastrin level and complete regression of initial low-grade MALT lymphoma was observed in all patients. However, two patients subsequently developed recurrent high-grade MALT lymphoma or high-grade lymphoma. In one of them, the serum gastrin level rose again above the pretreatment value. In the other, however, the fasting gastrin level fell throughout the study period. The median fasting serum gastrin level before H. pylori eradication therapy was higher in the patients with tumours of the gastric body (203.4 +/- 108.9 pg/ml) than in those with tumours of the antrum and angulus (89.3 +/- 28.0 pg/ml) (P = 0.06). CONCLUSIONS: Hypergastrinaemia may be associated with an increased risk of gastric MALT lymphoma.

Inhibition of CHOP translation by a peptide encoded by an open reading frame localized in the chop 5'UTR.

Chop is a ubiquitously expressed mammalian gene encoding a small nuclear protein related to the CCAAT/enhancer-binding protein (C/EBP) family of transcription factors. CHOP protein plays an important role in various cellular processes such as growth, differentiation and programmed cell death. CHOP expression is strongly increased in response to a large variety of stresses including perturbation of the endoplasmic reticulum function, DNA damage and nutrient deprivation. Multiple mechanisms including transcriptional and post-transcriptional controls are involved in the regulation of CHOP expression. We show here that the 5'UTR of the Chop transcript plays an important role in controlling the synthesis of CHOP protein. In particular, the 5'UTR contains a conserved uORF which encodes a 31 amino acid peptide that inhibits the expression of the downstream ORF. Mutational analysis of the 5' leader region and peptide coding sequences suggests that the peptide itself inhibits expression of the downstream ORF. Such results suggest a role for uORF in limiting ribosomal access to downstream initiation sites. With respect to the importance of CHOP protein in the regulation of cellular functions, the mechanisms that regulate its basal level are of considerable interest.

Catecholamines and serotonin are differently regulated by tetrahydrobiopterin. A study from 6-pyruvoyltetrahydropterin synthase knockout mice.

(6R)-L-erythro-5,6,7,8-Tetrahydrobiopterin (BH4) is an essential cofactor for tyrosine hydroxylase (TH), tryptophan hydroxylase, phenylalanine hydroxylase, and nitric-oxide synthase. These enzymes synthesize neurotransmitters, e.g. catecholamines, serotonin, and nitric oxide (NO). We established mice unable to synthesize BH4 by disruption of the 6-pyruvoyltetrahydropterin synthase gene, the encoded protein of which catalyzes the second step of BH4 biosynthesis. Homozygous mice were born at the almost expected Mendelian ratio, but died within 48 h after birth. In the brain of homozygous mutant neonates, levels of biopterin, catecholamines, and serotonin were extremely low. The number of TH molecules was highly dependent on the intracellular concentration of BH4 at nerve terminals. Alteration of the TH protein level by modulation of the BH4 content is a novel regulatory mechanism. Our data showing that catecholaminergic, serotonergic, and NO systems were differently affected by BH4 starvation suggest the possible involvement of BH4 synthesis in the etiology of monoamine-based neurological and neuropsychiatric disorders.

IRE1 and efferent signaling from the endoplasmic reticulum.

Genetic analysis of the cellular adaptation to malfolded proteins in the endoplasmic reticulum (the unfolded protein response - UPR) has revealed a novel signaling pathway initiated by activation of IRE1, an ER-resident protein kinase and endonuclease. In yeast, Ire1p activates gene expression by promoting a non-conventional splicing event that converts the mRNA encoding the Hac1p transcription factor from an inefficiently translated inactive mRNA to an actively translated one. Hac1p binds to the promoters of genes encoding chaperones and other targets of the UPR and activates them. Recently, mammalian IRE1 homologues have been identified and their response to ER stress is regulated by binding to the ER chaperone BiP. The mechanisms by which mammalian IRE1 activates gene expression have not been completely characterized and mammalian HAC1 homologues have not been identified. Surprisingly, mammalian IRE1s are able to activate both JUN N-terminal kinases and an alternative ER-stress signaling pathway mediated by the transcription factor ATF6. This indicates that the mammalian UPR is more complex than that found in yeast.

Triple transduction with adeno-associated virus vectors expressing tyrosine hydroxylase, aromatic-L-amino-acid decarboxylase, and GTP cyclohydrolase I for gene therapy of Parkinson's disease.

Parkinson's disease (PD), a neurological disease suited to gene therapy, is biochemically characterized by a severe decrease in the dopamine content of the striatum. One current strategy for gene therapy of PD involves local production of dopamine in the striatum achieved by inducing the expression of enzymes involved in the biosynthetic pathway for dopamine. We previously showed that the coexpression of tyrosine hydroxylase (TH) and aromatic-L-amino-acid decarboxylase (AADC), using two separate adeno-associated virus (AAV) vectors, resulted in more effective dopamine production and more remarkable behavioral recovery in 6-hydroxydopamine-lesioned parkinsonian rats, compared with the expression of TH alone. Not only levels of TH and AADC but also levels of tetrahydrobiopterin (BH4), a cofactor of TH, and GTP cyclohydrolase I (GCH), a rate-limiting enzymes for BH4 biosynthesis, are reduced in parkinsonian striatum. In the present study, we investigated whether transduction with separate AAV vectors expressing TH, AADC, and GCH was effective for gene therapy of PD. In vitro experiments showed that triple transduction with AAV-TH, AAV-AADC, and AAV-GCH resulted in greater dopamine production than double transduction with AAV-TH and AAV-AADC in 293 cells. Furthermore, triple transduction enhanced BH4 and dopamine production in denervated striatum of parkinsonian rats and improved the rotational behavior of the rats more efficiently than did double transduction. Behavioral recovery persisted for at least 12 months after stereotaxic intrastriatal injection. These results suggest that GCH, in addition to TH and AADC, is important for effective gene therapy of PD.

Male sterility and enhanced radiation sensitivity in TLS(-/-) mice.

TLS (also known as FUS) is an RNA-binding protein that contributes the N-terminal half of fusion oncoproteins implicated in the development of human liposarcomas and leukemias. Here we report that male mice homozygous for an induced mutation in TLS are sterile with a marked increase in the number of unpaired and mispaired chromosomal axes in pre-meiotic spermatocytes. Nuclear extracts from TLS(-/-) testes lack an activity capable of promoting pairing between homologous DNA sequences in vitro, and TLS(-/-) mice and embryonic fibroblasts exhibit increased sensitivity to ionizing irradiation. These results are consistent with a role for TLS in homologous DNA pairing and recombination.

Acute myeloid leukemia possessing jumping translocation is related to highly elevated levels of EAT/mcl-1, a Bcl-2 related gene with anti-apoptotic functions.

Jumping translocations (JTs) are unbalanced chromosomal translocations in which an identical chromosomal region is translocated to the telomeric region of different chromosomes. JTs are rare in hematological malignancies where they are second translocations and may be an indicator of poor prognosis. We report a case of acute myeloid leukemia with t(16;21) and a JT in which the long arm of chromosome 1 distal to q21 is translocated to the terminal region of chromosome 10. The leukemic cells exhibit high expression of EAT/mcl1, an anti-apoptotic Bcl-2 related gene. Since EAT/mcl1 is mapped to 1q21 near the breakpoint in the JTs, high level expression of EAT/mcl1 may be associated with the poor prognosis of leukemia with JTs.

Coupling of stress in the ER to activation of JNK protein kinases by transmembrane protein kinase IRE1.

Malfolded proteins in the endoplasmic reticulum (ER) induce cellular stress and activate c-Jun amino-terminal kinases (JNKs or SAPKs). Mammalian homologs of yeast IRE1, which activate chaperone genes in response to ER stress, also activated JNK, and IRE1alpha-/- fibroblasts were impaired in JNK activation by ER stress. The cytoplasmic part of IRE1 bound TRAF2, an adaptor protein that couples plasma membrane receptors to JNK activation. Dominant-negative TRAF2 inhibited activation of JNK by IRE1. Activation of JNK by endogenous signals initiated in the ER proceeds by a pathway similar to that initiated by cell surface receptors in response to extracellular signals.

Upregulation of BiP and CHOP by the unfolded-protein response is independent of presenilin expression.

Presenilin 1 (PS1), a polytopic membrane protein, has a critical role in the trafficking and proteolysis of a selected set of transmembrane proteins. The vast majority of individuals affected with early onset familial Alzheimer's disease (FAD) carry missense mutations in PS1. Two studies have suggested that loss of PS1 function, or expression of FAD-linked PS1 variants, compromises the mammalian unfolded-protein response (UPR), and we sought to evaluate the potential role of PS1 in the mammalian UPR. Here we show that that neither the endoplasmic reticulum (ER) stress-induced accumulation of BiP and CHOP messenger RNA, nor the activation of ER stress kinases IRE1alpha and PERK, is compromised in cells lacking both PS1 and PS2 or in cells expressing FAD-linked PS1 variants. We also show that the levels of BiP are not significantly different in the brains of individuals with sporadic Alzheimer's disease or PS1-mediated FAD to levels in control brains. Our findings provide evidence that neither loss of PS1 and PS2 function, nor expression of PS1 variants, has a discernable impact on ER stress-mediated induction of the several established 'readouts' of the UPR pathway.