研究者詳細 - 上原　孝

論文 - 上原　孝

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Reciprocal regulation of ATPgammaS-induced monocyte chemoattractant protein-1 production by ERK and p38 MAP kinases in rat corticostriatal slice cultures. 国際誌

Takahiro Katayama, Misato Ito, Shuji Kaneko, Masamichi Satoh, Takashi Uehara, Masabumi Minami

Journal of neuroscience research 87 ( 7 ) 1573 - 81 2009年5月

記述言語：英語掲載種別：研究論文（学術雑誌）

Monocyte chemoattractant protein-1 (MCP-1, CCL2) is a well-defined chemokine implicated in the pathology of various neurodegenerative diseases and brain injuries, such as Alzheimer's disease, multiple sclerosis, stroke, and traumatic injury. We investigated the effect of the activation of P2 purinoceptors on MCP-1 production in rat corticostriatal slice cultures. Treatment with adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS), a hydrolysis-resistant adenosine triphosphate (ATP) analog, induced MCP-1 production in astrocytes. The induction was in a concentration-dependent manner and was antagonized by a P2 purinoceptor antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid. The inhibition of an extracellular signal-regulated kinase (ERK) pathway by PD98059 and U0126 significantly suppressed ATPgammaS-induced MCP-1 mRNA expression and protein production, while inhibition of c-Jun N-terminal kinase by SP600125 resulted in the partial suppression. Conversely, SB203580, a p38 mitogen-activated protein (MAP) kinase inhibitor, significantly enhanced ATPgammaS-induced MCP-1 production. Similar effects of ERK and p38 MAP kinase inhibitors on MCP-1 production were observed in the slices stimulated by ATP and BzATP. These results demonstrate that astrocytic MCP-1 production induced by P2 purinoceptor stimulation is reciprocally regulated by ERK and p38 MAP kinases in the organotypic slice cultures.

DOI： 10.1002/jnr.21982

Neuronal injury induces cytokine-induced neutrophil chemoattractant-1 (CINC-1) production in astrocytes.

Takahiro Katayama, Hiroki Tanaka, Tadashi Yoshida, Takashi Uehara, Masabumi Minami

Journal of pharmacological sciences 109 ( 1 ) 88 - 93 2009年1月

記述言語：英語掲載種別：研究論文（学術雑誌）

Accumulating evidence indicates a pivotal role for neuroinflammation in ischemic and excitotoxic brain injury. Cytokine-induced neutrophil chemoattractant-1 (CINC-1) is a CXC chemokine implicated in the infiltration of inflammatory cells into the brain parenchyma. In this study, we investigated the effect of N-methyl-D-aspartate (NMDA)-induced neuronal injury on CINC-1 production in the organotypic cortico-striatal slice cultures. Treatment with 50 microM NMDA for 3 - 4 h caused devastating neuronal damage and increased CINC-1 production. Immunohistochemical analysis revealed that the CINC-1 immunoreactivity was predominantly detected in astrocytes. NMDA failed to induce CINC-1 production in enriched astrocyte cultures or neuron-depleted slice cultures, suggesting that NMDA acted on neuronal cells to induce astrocytic CINC-1 production. NMDA-induced CINC-1 mRNA expression was significantly inhibited by U0126, a mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor. These results suggest that NMDA-evoked neuronal injury induced astrocytic CINC-1 production via a MEK/ERK signaling pathway. Manipulation of this signaling pathway may serve as a target for suppressing neuroinflammation and, thereby, treating ischemic brain injury.

GMEB1, a novel endogenous caspase inhibitor, prevents hypoxia- and oxidative stress-induced neuronal apoptosis. 国際誌

Tadashi Nakagawa, Kazuhiro Tsuruma, Takashi Uehara, Yasuyuki Nomura

Neuroscience letters 438 ( 1 ) 34 - 7 2008年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

The interaction of glucocorticoid modulatory element-binding protein 1 (GMEB1) with procaspase-2, -8, or -9 prevents caspase oligomerization and maturation. In the present study, we examined the effect of GMEB1 on neuronal apoptosis induced by hypoxia and oxidative stress. GMEB1 effectively attenuated caspase activation and apoptosis caused by these stresses in human neuroblastoma SK-N-MC cells, indicating that it functions as a potent inhibitor of caspase activation and apoptosis in response to oxidative stress. We propose that GMEB1 blocks pro-apoptosis signals induced by a variety of stresses.

DOI： 10.1016/j.neulet.2008.04.023

Accumulation of misfolded protein through nitrosative stress linked to neurodegenerative disorders. 国際誌

Takashi Uehara

Antioxidants & redox signaling 9 ( 5 ) 597 - 601 2007年5月

記述言語：英語掲載種別：研究論文（学術雑誌）

Protein quality control is a critical feature of intracellular homeostasis. In particular, unfolded or misfolded proteins resulting from environmental stresses or free radicals are rapidly degraded via the ubiquitin-proteasome pathway. Nitric oxide (NO), a free radical gas, has been reported to be involved in such processes as vasorelaxation and neurotransmission. Conversely, NO also is implicated in neuronal cell death or neurodegeneration. Recent reports suggest that S-nitrosylation of proteins is a significant cause of neural dysfunction leading to neurodegenerative disorders. Specifically, S-nitrosylation of parkin eventually leads to the accumulation of unfolded proteins and subsequent neuronal death. The focus of this review is the identity of the target of NO. Nitrosative stress prevents normal functioning of the endoplasmic reticulum (ER) via S-nitrosylation of protein-disulfide isomerase (PDI), which is located in the ER lumen. This may contribute to the accumulation of misfolded proteins, as well as sustained activation of the unfolded protein response (UPR) pathway. These phenomena may be linked to the development of sporadic neurodegenerative diseases.

Involvement of the bed nucleus of the stria terminalis in the negative affective component of visceral and somatic pain in rats. 国際誌

Satoshi Deyama, Takayuki Nakagawa, Shuji Kaneko, Takashi Uehara, Masabumi Minami

Behavioural brain research 176 ( 2 ) 367 - 71 2007年1月

記述言語：英語掲載種別：研究論文（学術雑誌）

Using a conditioned place paradigm, we examined the involvement of the bed nucleus of the stria terminalis (BST) in the negative affective component of visceral and somatic pain induced by intraperitoneal acetic acid and intraplantar formalin injections, respectively, in rats. Bilateral BST lesions suppressed both the acetic acid- and formalin-induced conditioned place aversion, suggesting the crucial role of the BST in the negative affective component of visceral and somatic pain.

A ubiquitin ligase HRD1 promotes the degradation of Pael receptor, a substrate of Parkin. 国際誌

Tomohiro Omura, Masayuki Kaneko, Yasunobu Okuma, Yasuko Orba, Kazuo Nagashima, Ryosuke Takahashi, Noboru Fujitani, Satoshi Matsumura, Akihisa Hata, Kyoko Kubota, Karin Murahashi, Takashi Uehara, Yasuyuki Nomura

Journal of neurochemistry 99 ( 6 ) 1456 - 69 2006年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

It has been proposed that in autosomal recessive juvenile parkinsonism (AR-JP), a ubiquitin ligase (E3) Parkin, which is involved in endoplasmic reticulum-associated degradation (ERAD), lacks E3 activity. The resulting accumulation of Parkin-associated endothelin receptor-like receptor (Pael-R), a substrate of Parkin, leads to endoplasmic reticulum stress, causing neuronal death. We previously reported that human E3 HRD1 in the endoplasmic reticulum protects against endoplasmic reticulum stress-induced apoptosis. This study shows that HRD1 was expressed in substantia nigra pars compacta (SNC) dopaminergic neurons and interacted with Pael-R through the HRD1 proline-rich region, promoting the ubiquitylation and degradation of Pael-R. Furthermore, the disruption of endogenous HRD1 by small interfering RNA (siRNA) induced Pael-R accumulation and caspase-3 activation. We also found that ATF6 overexpression, which induced HRD1, accelerated and caused Pael-R degradation; the suppression of HRD1 expression by siRNA partially prevents this degradation. These results suggest that in addition to Parkin, HRD1 is also involved in the degradation of Pael-R.

Suppressive effects of 4-phenylbutyrate on the aggregation of Pael receptors and endoplasmic reticulum stress. 国際誌

Kyoko Kubota, Yoshifumi Niinuma, Masayuki Kaneko, Yasunobu Okuma, Mami Sugai, Tomohiro Omura, Mai Uesugi, Takashi Uehara, Toru Hosoi, Yasuyuki Nomura

Journal of neurochemistry 97 ( 5 ) 1259 - 68 2006年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

Endoplasmic reticulum (ER) stress is defined as an accumulation of unfolded proteins in the endoplasmic reticulum. 4-phenylbutyrate (4-PBA) has been demonstrated to promote the normal trafficking of the DeltaF508 cystic fibrosis transmembrane conductance regulator (CFTR) mutant from the ER to the plasma membrane and to restore activity. We have reported that 4-PBA protected against cerebral ischemic injury and ER stress-induced neuronal cell death. In this study, we revealed that 4-PBA possesses chemical chaperone activity in vitro, which prevents the aggregation of denatured alpha-lactalbumin and bovine serum albumin (BSA). Furthermore, we investigated the effects of 4-PBA on the accumulation of Parkin-associated endothelin receptor-like receptor (Pael-R) pathologically relevant to the loss of dopaminergic neurons in autosomal recessive juvenile parkinsonism (AR-JP). Interestingly, 4-PBA restored the normal expression of Pael-R protein and suppressed ER stress induced by the overexpression of Pael-R. In addition, we showed that 4-PBA attenuated the activation of ER stress-induced signal transduction pathways and subsequent neuronal cell death. Moreover, 4-PBA restored the viability of yeasts that fail to induce an ER stress response under ER stress conditions. These results suggest that 4-PBA suppresses ER stress by directly reducing the amount of misfolded protein, including Pael-R accumulated in the ER.

S-nitrosylated protein-disulphide isomerase links protein misfolding to neurodegeneration. 国際誌

Takashi Uehara, Tomohiro Nakamura, Dongdong Yao, Zhong-Qing Shi, Zezong Gu, Yuliang Ma, Eliezer Masliah, Yasuyuki Nomura, Stuart A Lipton

Nature 441 ( 7092 ) 513 - 7 2006年5月

記述言語：英語掲載種別：研究論文（学術雑誌）

Stress proteins located in the cytosol or endoplasmic reticulum (ER) maintain cell homeostasis and afford tolerance to severe insults. In neurodegenerative diseases, several chaperones ameliorate the accumulation of misfolded proteins triggered by oxidative or nitrosative stress, or of mutated gene products. Although severe ER stress can induce apoptosis, the ER withstands relatively mild insults through the expression of stress proteins or chaperones such as glucose-regulated protein (GRP) and protein-disulphide isomerase (PDI), which assist in the maturation and transport of unfolded secretory proteins. PDI catalyses thiol-disulphide exchange, thus facilitating disulphide bond formation and rearrangement reactions. PDI has two domains that function as independent active sites with homology to the small, redox-active protein thioredoxin. During neurodegenerative disorders and cerebral ischaemia, the accumulation of immature and denatured proteins results in ER dysfunction, but the upregulation of PDI represents an adaptive response to protect neuronal cells. Here we show, in brains manifesting sporadic Parkinson's or Alzheimer's disease, that PDI is S-nitrosylated, a reaction transferring a nitric oxide (NO) group to a critical cysteine thiol to affect protein function. NO-induced S-nitrosylation of PDI inhibits its enzymatic activity, leads to the accumulation of polyubiquitinated proteins, and activates the unfolded protein response. S-nitrosylation also abrogates PDI-mediated attenuation of neuronal cell death triggered by ER stress, misfolded proteins or proteasome inhibition. Thus, PDI prevents neurotoxicity associated with ER stress and protein misfolding, but NO blocks this protective effect in neurodegenerative disorders through the S-nitrosylation of PDI.

Glucocorticoid modulatory element-binding protein 1 binds to initiator procaspases and inhibits ischemia-induced apoptosis and neuronal injury. 国際誌

Kazuhiro Tsuruma, Tadashi Nakagawa, Nobutaka Morimoto, Masabumi Minami, Hideaki Hara, Takashi Uehara, Yasuyuki Nomura

The Journal of biological chemistry 281 ( 16 ) 11397 - 404 2006年4月

記述言語：英語掲載種別：研究論文（学術雑誌）

Caspases are divided into two classes: initiator caspases, which include caspase-8 and -9 and possess long prodomains, and effector caspases, which include caspase-3 and -7 and possess short prodomains. Recently, we demonstrated that glucocorticoid modulatory element-binding protein 1 (GMEB1) interacts with the prodomain of procaspase-2, thereby disrupting its autoactivation and the induction of apoptosis. Here we show that GMEB1 is also capable of binding to procaspase-8 and -9. GMEB1 attenuated the Fas-mediated activation of these caspases and the subsequent apoptosis. The knockdown of endogenous GMEB1 using RNA interference revealed that cells with decreased GMEB1 expression are more sensitive to stress and undergo accelerated apoptosis. Transgenic mice expressing a neurospecific GMEB1 had smaller cerebral infarcts and less brain swelling than wild-type mice in response to transient focal ischemia. These results suggest that GMEB1 is an endogenous regulator that selectively binds to initiator procaspases and inhibits caspase-induced apoptosis.

Astrapterocarpan isolated from Astragalus membranaceus inhibits proliferation of vascular smooth muscle cells. 国際誌

Susumu Ohkawara, Yasunobu Okuma, Takashi Uehara, Takashi Yamagishi, Yasuyuki Nomura

European journal of pharmacology 525 ( 1-3 ) 41 - 7 2005年11月

記述言語：英語掲載種別：研究論文（学術雑誌）

The inhibitory effects of astrapterocarpan, formononetin, and calycosin isolated from Astragalus membraneceus on platelet-derived growth factor (PDGF)-BB-induced proliferative response in rat vascular smooth muscle cells (A10 cells) were investigated. Astrapterocarpan significantly inhibited PDGF-BB-induced cell proliferation and DNA synthesis in a concentration-dependent manner. This inhibition was not attributed to toxicity. In contrast, formononetin and calycosin had no effect. We next examined the effect of astrapterocarpan on PDGF-BB signal transduction. Astrapterocarpan inhibited PDGF-BB-induced phosphorylation of extracellular signal-regulated kinase 1/2 (ERIC1/2) mitogen-activated protein (MAP) kinase. However, this compound had no effect on phosphorylation of PDGF-beta-receptor, Akt kinase and p38 MAP kinase. These results indicated that astrapterocarpan inhibits PDGF-BB-induced vascular smooth muscle cell proliferation and that this effect may be mediated, at least in part, by inhibition of the ERK1/2 MAP kinase cascade.

Comment on "S-nitrosylation of parkin regulates ubiquitination and compromises parkin's protective function". 国際誌

Stuart A Lipton, Tomohiro Nakamura, Dongdong Yao, Zhong-Qing Shi, Takashi Uehara, Zezong Gu

Science (New York, N.Y.) 308 ( 5730 ) 1870; author reply 1870 2005年6月

Caspase recruitment domain of procaspase-2 could be a target for SUMO-1 modification through Ubc9. 国際誌

Hiromi Shirakura, Naoko Hayashi, Shin-ichi Ogino, Kazuhiro Tsuruma, Takashi Uehara, Yasuyuki Nomura

Biochemical and biophysical research communications 331 ( 4 ) 1007 - 15 2005年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

To identify the binding proteins that regulate the function of procaspase-2, we screened for proteins using the yeast two-hybrid method and isolated human Ubc9 and SUMO-1 as the candidates. Ubc9 and SUMO-1 interacted with the caspase recruitment domain of procaspase-2 in its N-terminal. We elucidated the covalent modification of procaspase-2 by SUMO-1 in mammalian cells by immunoprecipitation followed by Western blot analysis. Procaspase-2 and SUMO-1 were co-localized by dot-like structures in the nucleus that are related to promyelocytic leukemia bodies. Interestingly, a conjugation-deficient mutant (K60R) procaspase-2 resulted in a delay of its enzyme maturation (appearance of p12 subunit) compared to that of wild-type. Thus, the modification with SUMO-1 may play a critical role in the nuclear localization and the activation (maturation) of procaspase-2.

Regulation of procaspase-2 by glucocorticoid modulatory element-binding protein 1 through the interaction with caspase recruitment domain. 国際誌

Kazuhiro Tsuruma, Tadashi Nakagawa, Hiromi Shirakura, Naoko Hayashi, Takashi Uehara, Yasuyuki Nomura

Biochemical and biophysical research communications 325 ( 4 ) 1246 - 51 2004年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

Caspases are the primary executioners of apoptosis. Although procaspases believe to exist as inactive forms in cells, the detailed regulatory system remains unclear. Here we show that glucocorticoid modulatory element-binding protein 1 (GMEB1) is capable of binding to the prodomain of caspase-2. We found that this molecule inhibits the autoproteolytic activation of procaspase-2 by oligomerization on a chemical compound-dependent system. These findings indicated that GMEB1 might be an endogenous inhibitory protein that selectively interacts with prodomain of caspase-2 to disrupt the autoactivation.

Possible involvement of pyruvate kinase in acquisition of tolerance to hypoxic stress in glial cells. 国際誌

Takahiro Shimizu, Takashi Uehara, Yasuyuki Nomura

Journal of neurochemistry 91 ( 1 ) 167 - 75 2004年10月

記述言語：英語掲載種別：研究論文（学術雑誌）

Neurons are highly vulnerable to ischemic/hypoxic stress, while glial cells show tolerance to such stress. However, the mechanisms for tolerance acquisition in glial cells have yet to be established. We attempted to isolate and identify a stress protein that is upregulated in response to hypoxia in human astrocytoma CCF-STTG1 cells. In particular, pyruvate kinase (PK) was upregulated by hypoxia in CCF-STTG1 cells. Hypoxia-inducible factor 1 (HIF-1), the primary transcription factor that is responsible for multiple gene activation under hypoxia, plays a critical role in PK expression during hypoxic challenge. To determine whether newly synthesized PK is involved in tolerance to hypoxic stress, we established the PK-overexpressing neuronal cells. Overexpression of the wild-type, but not the kinase-negative mutant, resulted in attenuation of the loss of cell viability and the typical apoptotic features by hypoxia or oxidative stress in SK-N-MC cells. These findings suggest that upregulation of PK may result in acquisition of tolerance against hypoxic stress, and that the antioxidant effect may be involved in the protective effect of PK.

Nitrosative stress linked to sporadic Parkinson's disease: S-nitrosylation of parkin regulates its E3 ubiquitin ligase activity. 国際誌

Dongdong Yao, Zezong Gu, Tomohiro Nakamura, Zhong-Qing Shi, Yuliang Ma, Benjamin Gaston, Lisa A Palmer, Edward M Rockenstein, Zhuohua Zhang, Eliezer Masliah, Takashi Uehara, Stuart A Lipton

Proceedings of the National Academy of Sciences of the United States of America 101 ( 29 ) 10810 - 4 2004年7月

記述言語：英語掲載種別：研究論文（学術雑誌）

Many hereditary and sporadic neurodegenerative disorders are characterized by the accumulation of aberrant proteins. In sporadic Parkinson's disease, representing the most prevalent movement disorder, oxidative and nitrosative stress are believed to contribute to disease pathogenesis, but the exact molecular basis for protein aggregation remains unclear. In the case of autosomal recessive-juvenile Parkinsonism, mutation in the E3 ubiquitin ligase protein parkin is linked to death of dopaminergic neurons. Here we show both in vitro and in vivo that nitrosative stress leads to S-nitrosylation of wild-type parkin and, initially, to a dramatic increase followed by a decrease in the E3 ligase-ubiquitin-proteasome degradative pathway. The initial increase in parkin's E3 ubiquitin ligase activity leads to autoubiquitination of parkin and subsequent inhibition of its activity, which would impair ubiquitination and clearance of parkin substrates. These findings may thus provide a molecular link between free radical toxicity and protein accumulation in sporadic Parkinson's disease.

Herbimycin A abrogates nuclear factor-kappaB activation by interacting preferentially with the IkappaB kinase beta subunit. 国際誌

Shinichi Ogino, Kazuhiro Tsuruma, Takashi Uehara, Yasuyuki Nomura

Molecular pharmacology 65 ( 6 ) 1344 - 51 2004年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

NF (nuclear factor)-kappaB is known to be a critical transcription factor in inflammatory responses. We have reported that herbimycin A, a potent Src tyrosine kinase inhibitor, attenuates the NF-kappaB activation triggered by cytokines, bacterial endotoxin, and hydrogen peroxide. Accompanying the suppression by this agent, NF-kappaB-dependent gene expressions, such as cytokine, chemokine, and inducible-type nitric oxide, are specifically inhibited in glial cells. In the present study, we attempted to elucidate the possible target protein for herbimycin A on this pathway. We demonstrate here that herbimycin A preferentially inhibits IKK (IkappaB kinase)beta. Furthermore, substituting alanine for the cysteine at 59 (Cys59) in IKKbeta resulted in the insensitivity to herbimycin A, suggesting that this compound may interact with the Cys59 residue located near the catalytic ATP binding site. Taken together, these results indicate that herbimycin A can be considered a novel candidate for an anti-inflammatory drug agent through its specific inhibition of IKKbeta, which results in prevention of the expression of NF-kappaB-dependent genes implicated in the pathogenesis of inflammatory responses.

Ubiquilin interacts with ubiquitylated proteins and proteasome through its ubiquitin-associated and ubiquitin-like domains. 国際誌

Han Seok Ko, Takashi Uehara, Kazuhiro Tsuruma, Yasuyuki Nomura

FEBS letters 566 ( 1-3 ) 110 - 4 2004年5月

記述言語：英語掲載種別：研究論文（学術雑誌）

Mammalian cells acquire tolerance against multiple stressors through the high-level expression of stress-responsible genes. We have previously demonstrated that protein-disulfide isomerase (PDI) together with ubiquilin are up-regulated in response to hypoxia/brain ischemia, and play critical roles in resistance to these damages. We show here that ubiquilin interacts preferentially with poly-ubiquitin chains and 19S proteasome subunits. Taken together, these results suggest that ubiquitin could serve as an adaptor protein that both interacts with PDI and mediates the delivery of poly-ubiquitylated proteins to the proteasome in the cytosol in the vicinity of the endoplasmic reticulum membrane.

Correlation between delayed neuronal cell death and selective decrease in phosphatidylinositol 4-kinase expression in the CA1 subfield of the hippocampus after transient forebrain ischemia. 国際誌

Yudai Furuta, Takashi Uehara, Yasuyuki Nomura

Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism 23 ( 8 ) 962 - 71 2003年8月

記述言語：英語掲載種別：研究論文（学術雑誌）

Transient forebrain ischemia induces a delayed neuronal death in the CA1 area of the hippocampus. However, the mechanism leading to this phenomenon has yet to be established. The authors used an mRNA differential-display method to isolate genes for which mRNA levels change only in the hippocampus during ischemia/reperfusion. They succeeded in identifying the product of one down-regulated gene as phosphatidylinositol 4-kinase (PI 4-K). Compared with control levels, PI 4-K mRNA expression in the hippocampus, but not the cerebral cortex, was significantly decreased by 30% and about 80% 1 and 7 days after ischemia/reperfusion, respectively. Interestingly, PI 4-K and PI bisphosphate levels were selectively decreased in the CA1 region, but not other regions, whereas TUNEL-positive cells could be detected 3 days after ischemia. Consistent with these results, PI 4-K expression was suppressed by hypoxia in SK-N-MC neuroblastoma cells before loss of cell viability. Overexpression of wild-type PI 4-K, but not the kinase-negative mutant of PI 4-K (K1789A), recovered the loss of viability induced by hypoxia. These findings strongly suggest that a prior decrease in PI 4-K and PI bisphosphate levels caused by brain ischemia/hypoxia is partly involved in delayed neuronal cell death.

Role of ubiquilin associated with protein-disulfide isomerase in the endoplasmic reticulum in stress-induced apoptotic cell death. 国際誌

Han Seok Ko, Takashi Uehara, Yasuyuki Nomura

The Journal of biological chemistry 277 ( 38 ) 35386 - 92 2002年9月

記述言語：英語掲載種別：研究論文（学術雑誌）

Up-regulation of several stress proteins such as heat-shock proteins and glucose-regulated proteins participate in tolerance against environmental stress. Previously, we found that protein-disulfide isomerase (PDI) is specifically up-regulated in response to hypoxia/brain ischemia in astrocytes. In addition, the overexpression of this gene into neurons protects against apoptotic cell death induced by hypoxia/brain ischemia. To address the detailed function of PDI, we screened for proteins that interact with PDI using the yeast two-hybrid system. We report here that PDI interacts with ubiquilin, which has a ubiquitin-like domain and a ubiquitin-associated domain. Interestingly, ubiquilin is also up-regulated in response to hypoxia in glial cells with a time course similar to that of PDI induction. In hypoxia-treated glial cells, the endogenous ubiquilin and PDI were almost completely co-localized, suggesting that ubiquilin is an endoplasmic reticulum-associated protein. Overexpression of this gene in neuronal cells resulted in significant inhibition of the DNA fragmentation triggered by hypoxia, but not that induced by nitric oxide or staurosporine. Moreover, ubiquilin has the ability to attenuate CHOP induction by hypoxia. These observations suggested that ubiquilin together with PDI have critical functions as regulatory proteins for CHOP-mediated cell death, and therefore up-regulation of these proteins may result in acquisition of tolerance against ischemic stress in glial cells.

Involvement of nuclear factor-kappa B (NF-kappa B) signaling in the expression of inducible nitric oxide synthase (iNOS) gene in rat C6 glioma cells 査読

T Nishiya, T Uehara, M Kaneko, Y Nomura

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 275 ( 2 ) 268 - 273 2000年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC

It has been demonstrated from studies using NF-kappa B inhibitors that NF-kappa B may be involved in the iNOS induction stimulated by cytokines and/or lipopolysaccharide (LPS) in various cell types and tissues. However, the actions of the inhibitors are less selective and highly cytotoxic, We constructed stable clones of C6 cells transfected with two types of I kappa B alpha mutant genes (I kappa B alpha(SS-->AA); Ser-32/36 to Ala-32/36, I kappa B alpha(KK-->RR);Lys-21/22 to Arg-21/22). I kappa B alpha(SS-->AA) strongly inhibited (1) LPS-, IL-1 beta-, and TNF-alpha-induced nuclear translocation and DNA binding of NF-kappa B to the kappa B site; and (2) iNOS induction stimulated by LPS or IL-1 beta plus IFN-gamma, These results indicate that NF-kappa B plays a critical role in cytokines and/or LPS-induced iNOS induction. Surprisingly, similar to the endogenous I kappa B alpha, I kappa B alpha(KK-->RR) was degraded by various stimuli, and proteasome inhibitors blocked this event. These results suggest that another Lys residue(s), other than Lys-21/22, may be required for the ligand-induced I kappa B alpha degradation by the ubiquitin-proteasome pathway. (C) 2000 Academic Press.

DOI： 10.1006/bbrc.2000.3293

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