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Volume 136 (2004)
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Volume 136, Issue 3
Displaying 1-20 of 20 articles from this issue
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Jun Matsukawa, Atsushi Matsuzawa, Kohsuke Takeda, Hidenori Ichijo2004 Volume 136 Issue 3 Pages 261-265
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe mitogen-activated protein (MAP) kinase cascades play essential roles in a variety of cell processes by influencing transcriptional or translational regulation. ERKs play a central role in survival and mitogenic signaling, while JNKs and p 38 MAP kinases are preferentially activated by environmental stresses and are actively involved in various stress responses including cell death, survival and differentiation. Apoptosis signal-regulating kinase 1 (ASK1)-a serine/threonine protein kinase-is a member of the MAPKKK family and activates both JNK and p 38 path-ways. It is well known that ASK1 is activated in cells treated with death receptor ligands and oxidant stress, such as that caused by hydrogen peroxide (H2O2). Moreover, recent studies have revealed new mechanisms by which ASK1 is activated in response to various types of extracellular and intracellular signals, such as endoplasmic reticulum (ER) stress, calcium signaling, and G-protein coupled receptor (GPCR) signaling. This review summarizes the regulatory mechanisms of ASK1 activity and the physiological roles of ASK1-mediated signal transduction.View full abstractDownload PDF (1924K) -
Haruo Saito, Kazuo Tatebayashi2004 Volume 136 Issue 3 Pages 267-272
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe budding yeast Saccharomyces cerevisiae has at least five signal pathways containing a MAP kinase (MAPK) cascade. The high osmolarity glycerol (HOG) MAPK pathway is essential for yeast survival in high osmolarity environment. This mini-review surveys recent developments in regulation of the HOG pathway with specific emphasis on the roles of protein phosphatases and protein subcellular localization. The Hogl MAPK in the HOG pathway is negatively regulated jointly by the protein tyrosine phosphatases Ptp2/Ptp3 and the type 2 protein phosphatases Ptcl/Ptc2/Ptc3. Specificities of these phosphatases are determined by docking interactions as well as their cellular localizations. The subcellular localizations of the osmosensors (Slnl and Shol), kinases (Pbs2, Hogl), and phosphatases in the HOG pathway are intri-cately regulated to achieve their specific functions.View full abstractDownload PDF (2143K) -
Masashi Fukasawal, Terumasa Tsuchiya, Eiji Takayama, Nariyoshi Shinomi ...2004 Volume 136 Issue 3 Pages 273-277
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe placenta-type isozyme of human 6-phosphofructo-2-kinase/fructose-2, 6-bisphos-phatase (HP2K, identical to PFKFB3) is expressed in a variety of cells and tissues such as placenta, brain, testis, liver, kidney, skeletal muscle, primary blood mono-nuclear cells and cancer cells. We observed previously that the enhancer region of the HP2K gene, which has been identified in the 5'-flanking region between -1265 and -1329, could respond to serum stimulation following the transfection of human chori-ocarcinoma BeWo cells with HP2K promoter-luciferase constructs. The HP2K enhancer region also contains two copies of the hypoxia-inducible facif (5'-ACGTG-3'). In this study we performed characterization of the HP2K tor-1 (HIF-1) binding motgene expression in response to hypoxic conditions. Both electrophoretic mobility shift and co-transfection assays of the HP2K promoter-luciferase reporter with HIF-1 expression vectors indicated that HIF-1 binds to the hypoxia-responsive element (HRE) of HP2K, thereby upregulating its gene expression. In addition, we demon-strated using site-directed mutagenesis that a complete tandem repeat of the HIF-1 binding motif with a 4-bp interruption is required for full induction of HP2K expres-sion (up to 22-fold) under hypoxic conditions, and that this response is much stronger than that of the erythropoietin (EPO) gene. These results suggest that the sequence 5'-ACGTGNNNNACGTG-3' in the HP2K enhancer is the authentic HRE consensus motif that mediates increased transcription, under hypoxic conditions, via HIF-1.View full abstractDownload PDF (1985K) -
Satoshi Kubota, Kazumi Kawata, Takeshi Yanagita, Hideyuki Doi, Takashi ...2004 Volume 136 Issue 3 Pages 279-282
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSWound healing and tissue regeneration are usually initiated by coagulation followed by fibrous tissue formation. In the present study, we discovered an abundance of connective tissue growth factor (CTGF/CCN2) in human platelets, which was released along with the coagulation process. The CTGF/CCN2 content in platelets was 10-fold higher than that in arterial tissue. Furthermore, the CTGF/CCN2 content in a single platelet was computed to be more than 20-fold higher than that of any other growth factor reported. Considering that CTGF/CCN2 promotes angiogenesis, cartilage regeneration, fibrosis and platelet adhesion, it may be now regarded as one of the major functional components of platelets.View full abstractDownload PDF (1562K) -
Tatsuya Fukushima, Taichi Tanabe, Hiroki Yamamoto, Shigeo Hosoya, Tsut ...2004 Volume 136 Issue 3 Pages 283-291
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe predicted amino acid sequence of Bacillus subtilis ybaN (renamed pdaB) exhibits high similarity to those of several polysaccharide deacetylases. Northern hybridization analysis with sporulation sigma mutants indicated that the pdaB gene is tran-scribed by EδE RNA polymerase and negatively regulated by SpoIIID. The pdaB mutant was deficient in spore formation. Phase- and electron microscopic observation showed morphological changes of spores in late sporulation periods. The pdaB spores that had lost their viability were empty. Moreover, GFP driven by the promoter of the sspE gene was localized in the forespore compartment for the wild type, but was localized in both the mother cell and forespore compartments for phase-gray/dark forespores of the pdaB mutant. This indicates that GFP expressed in the fore-spores of the mutant leaks into the mother cells. Therefore, PdaB is necessary to maintain spores after the late stage of sporulation.View full abstractDownload PDF (6130K) -
Yukiko Umezawa, Keiichi Yokoyama, Yoshimi Kikuchi, Masayo Date, Kiyosh ...2004 Volume 136 Issue 3 Pages 293-300
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe prolyl peptidase that removes the tetra-peptide of pro-transglutaminase was purified from Streptomyces mobaraensis mycelia. The substrate specificity of the enzyme using synthetic peptide substrates showed proline-specific activity with not only tripeptidyl peptidase activity, but also tetrapeptidyl peptidase activity. However, the enzyme had no other exo- and endo-activities. This substrate specificity is different from proline specific peptidases so far reported. The enzyme gene was cloned, based on the direct N-terminal amino acid sequence of the purified enzyme, and the entire nucleotide sequence of the coding region was determined. The deduced amino acid sequence revealed an N-terminal signal peptide sequence (33 amino acids) followed by the mature protein comprising 444 amino acid residues. This enzyme shows no remarkable homology with enzymes belonging to the prolyl oligopeptidase family, but has about 65% identity with three tripeptidyl peptidases from Streptomyces lividans, Streptomyces coelicolor, and Streptomyces avermitilis. Based on its substrate specificity, a new name, “prolyl tri/tetra-peptidyl aminopeptidase, ” is proposed for the enzyme.View full abstractDownload PDF (3601K) -
Yoshihiro Matsumura, Tsuyoshi Kuzuguchi, Hiroshi Sagami2004 Volume 136 Issue 3 Pages 301-312
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSGeranylgeranyl diphosphate synthase (GGPS) is a branch point enzyme in the meval-onate pathway that catalyzes the synthesis of geranylgeranyl diphosphate used for the geranylgeranylation of Rho, Rac and Rab proteins. The current study showed the production of multiple forms of GGPS mRNA from a single GGPS gene in rat. The mRNAs resulted from combinations of multiple alternative introns and two poly (A) sites in the 3'-translated and 3'-untranslated regions. These are classified into 1 a-type and 1 b-type mRNAs, based on the splicing of intron 4 b resulting in the difference in deduced amino acid sequence between the C-terminal regions. The la-type and 1 b-type proteins expressed in both Escherichia coli and HeLa cells were active and inactive, respectively. In the case of HeLa cells, the latter protein expression level was about 10% relative to the former one. This was also observed for Cos-7 and 293 cells. When fusions of β-galactosidase with C-terminal regions differing between the latype and 1 b-type proteins were expressed in HeLa cells, the expressed fusion proteins were both found to be active but the latter fusion protein expression level was consid-erably low compared with the former one. The expression level of 1 a-type mRNA was higher than that of 1 b-type mRNA in brain, liver, heart, and thymus, but the two expression levels were the same in testis and ovary. During testis development the total GGPS mRNA expression level increased, accompanied by an increase in 1 b-type mRNA, the expression level of la-type mRNA encoding active GGPS remaining kept unchanged. These results indicate that the expression level of rat active GGPS is at least regulated through the splicing of intron 4 b of its gene.View full abstractDownload PDF (7206K) -
Takeaki Dohda, Hidenori Kaneoka, Yujin Inayoshi, Masamichi Kamihira, K ...2004 Volume 136 Issue 3 Pages 313-319
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSTranscriptional coactivators, CREB-binding protein (CBP) and p 300, exhibit high homology in structure and similar functions. In the present study, we analyzed the function of CBP and p 300 proteins as transcriptional coactivators in the expression of albumin in hepatocytes. The expression levels of CBP and p 300 were high in fetal hepatocytes, but low in adult ones. Immunoprecipitation assays showed that both CBP and p 300 interacted with hepatocyte nuclear factor-1 α (HNF-1 α) in primary hepatocytes. Furthermore, CBP and p 300 were co-precipitated without HNF-1 α, Chromatin immunoprecipitation (ChIP) assays revealed that both CBP and p 300 are located in the albumin promoter region in hepatocytes. These results suggested that HNF-1 α, CBP and p 300 were incorporated into a preinitiation complex of RNA polymerase II at the albumin promoter. Luciferase reporter assays showed that CBP and p 300 cooperatively triggered HNF-1 α-mediated transcription of the albumin pro-moter. In addition, inhibition of CBP or p 300 using small interfering RNAs (siRNAs) resulted in a reduction in albumin expression. These results suggest that both CBP and p 300 are required for enhanced expression of albumin.View full abstractDownload PDF (4752K) -
Shin Ebihara, Shota Endo, Kozue Ito, Yumi Ito, Kenichi Akiyama, Masuo ...2004 Volume 136 Issue 3 Pages 321-328
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe uptake of an antigen and its presentation to specific T cells by dendritic cells (DCs) is a primary event in initiation of humoral and cellular immune responses as well as the induction of cytotoxic T cells (CTLs). DCs are induced by culturing bone marrow cells in the presence of GM-CSF. However, the resulting DCs are short-lived and the culture usually contains CD11c-negative non-DC cells, which adversely affects reproducibility and makes interpretation of the experimental results difficult. Therefore, it would be useful if DCs could be readily immortalized with their functions being retained. In this study we established a novel, immortalized murine DC line with antigen-presenting capacity in vitro as well as an augmenting effect on humoral and cellular immune responses in vivo, utilizing bone marrow cells from transgenic mice harboring the temperature-sensitive SV40 large T-antigen gene. In the presence of GM-CSF, the resulting DC line, termed SVDC, could be continuously subcultured for more than 12 months. When pulsed with OVA alone or OVA-IgG immune complexes via Fcγ receptors, SVDC augmented OVA-specific T cell proliferation efficiently in vitro, and elicited OVA-specific IgG production in vivo on the adoptive transfer of pulsed SVDC into naive mice. Interestingly, SVDC exhibited significantly high cross-priming ability compared to DCs in a short-term culture, thus leading to their extremely high effectiveness in inducing anti-tumor immunity in vivo. Thus, SVDC is useful for the detailed characterization of antigen presentation, and for research on the various therapeutic benefits of DC vaccination to elicit specific immune responses in immunodeficiencies, infectious diseases and cancer.View full abstractDownload PDF (3321K) -
Arun Sharma, Alex Eapen, Sarala K. Subbarao2004 Volume 136 Issue 3 Pages 329-334
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSNitric oxide (NO) is known to possess antiparasitic activity towards Plasmodium species. Parasite proteases are currently considered to be promising targets for antima-larial chemotherapy. In the present study, we have studied the inhibitory effect of NO on the activity of plasmepsin in Plasmodium vivax, the pepsin-like aspartic protease which is believed to be involved in the cleavage during hemoglobin degradation in Plasmodium falciparum. NO donors (±) (E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (NOR-3), S-nitrosoglutathione (GSNO), and sodium nitroprusside (SNP) were found to inhibit this plasmepsin activity in a dose-dependent manner in purified P. vivax aspartic protease enzyme extracts. This inhibitory effect may be attribut-able to the nitrosylation of the cysteine residue at the catalytic site. However, an inhibitor of aspartic protease activity, namely pepstatin, was also found to inhibit (IC503 μM) the enzyme activity, which we have used as a positive control. Our results therefore provide novel insights into the pathophysiological mechanisms, and will be useful for designing strategies for selectively upregulating NO production in P. vivax infections for antimalarial chemotherapy and also biochemical adaptations of the malaria parasite for survival in the host erythrocytes with a better understanding of the protease substrate interactions.View full abstractDownload PDF (1821K) -
Keitarou Saiki, Tomoharu Gomi, Kiyoshi Konishi2004 Volume 136 Issue 3 Pages 335-342
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSCytolethal distending toxin (CDT) is one of the exotoxins produced by Actinobacillus actinomycetemcomitans, an agent of localized aggressive periodontitis. We constructed N-terminal deletion mutants of CdtA using an Escherichia coli expression system and found that AΔ19-47, with a deletion from Asn-19 to Pro-47, showed comparable CDT activity but no apparent heterogeneity of CdtA. The wild-type CDT (wtCDT) and the mutant CDT (AΔ19-47CDT) were purified to homogeneity by introducing a histidine tag into the C-terminal end of CdtB. Both purified wtCDT and purified AΔ19-47CDT showed strong CDT activity and a tripartite structure composed of CdtA (subunit A), 31 kDa CdtB (subunit B), and 18.5 kDa CdtC (subunit C) in nearly a 1:1:1 stoichiometry. Importantly, subunit A was identified as heterogeneous with three CdtA variants in wtCDT, but homogeneous in AΔ19-47CDT. Purified CDTs also showed high stability that was absolutely dependent on the presence of sucrose in the buffer. In conclusion, the region from the Asn-19 to Pro-47 of CdtA contributes to the hetero-geneous production of CdtA, but is dispensable for the toxin activity. Furthermore, this study describes an effective protocol for the purification of a native rather than reconstituted CDT, and clarifies the subunit composition of the active CDT holotoxin.View full abstractDownload PDF (4543K) -
Keisuke Yamamoto, Hiderou Yoshida, Koichi Kokame, Randal J. Kaufman, K ...2004 Volume 136 Issue 3 Pages 343-350
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSATF6 and XBP1 are transcription factors activated specifically in response to endo-plasmic reticulum (ER) stress. Three cis-acting elements capable of binding to ATF6, XBP1 or both have been identified to date, namely ER stress-response element (ERSE), unfolded protein response element (UPRE) and ERSE-II. ERSE controls the expression of ER-localized molecular chaperones such as BiP that can refold unfolded proteins in the ER; transcription from ERSE is fully activated by ATF6 even in the absence of XBP1. In contrast, transcription from UPRE depends solely on XBP1 and it has been suggested that UPRE may control the expression of components of the ER-associated degradation system that can degrade unfolded proteins in the ER. The Herp gene, one of the most highly inducible genes under ER stress, encodes an ER membrane protein containing a ubiquitin-like domain with unknown functions, and carries ERSE-II in addition to ERSE in its promoter. In this report, we show that ERSE-II allows the NF-Y-dependent binding of ATF6 as in the case of ERSE and NF-Y-independent binding of XBP1 as in the case of UPRE, and that transcription from ERSE-II is mitigated in the absence of XBP1. Accordingly, the induction of Herp mRNA was diminished in the absence of XBP1 whereas that of BiP mRNA was not affected. These results may help in understanding the role of Herp in the quality control system in the ER.View full abstractDownload PDF (4911K) -
Tadashi Jono, Ryoji Nagai, Xia Lin, Naila Ahmed, Paul J. Thornalley, M ...2004 Volume 136 Issue 3 Pages 351-358
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe levels of plasma 3-deoxyglucosone (3-DG) increase under hyperglycemic conditions and are associated with the pathogenesis of diabetic complications because of the high reactivity of 3-DG with proteins to form advanced glycation end products (AGE). To investigate potential markers for 3-DG-mediated protein modification in vitro and in vivo, we compared the yield of several 3-DG-derived AGE structures by immunochemical analysis and HPLC and measured their localization in human atherosclerotic lesions. When BSA was incubated with 3-DG at 37°C for up to 4 wk, the amounts of Nε-(carboxymethyl) lysine (CML) and 3-DG-imidazolone steeply increased with incubation time, whereas the levels of pyrraline and pentosidine increased slightly by day 28. In contrast, significants amount of pyrraline and pentosidine were also observed when BSA was incubated with 3-DG at 60°C to enhance AGE-formation. In atherosclerotic lesions, CML and 3-DG-imidazolone were found intracellularly in the cytoplasm of most foam cells and extracellularly in the atheromatous core. A weak-positive immunoreaction with pyrraline was found in the extracellular matrix and a few foam cells in aortic intima with atherosclerotic lesions. Our results provide the first evidence that CML and 3-DG-imidazolone are major AGE structures in 3-DG-modified proteins, and that 3-DG-imidazolone provides a better marker for protein modification by 3-DG than pyrraline.View full abstractDownload PDF (2360K) -
Hiroyuki Kamiya, Chieko Ishiguro, Hideyoshi Harashima2004 Volume 136 Issue 3 Pages 359-362
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe Escherichia coli MutT protein hydrolyzes 8-hydroxy-dGTP (8-OH-dGTP) in vitro, and mutT gene deficiencies cause increased spontaneous A: T→C: G mutations. However, no direct evidence exists for enhanced mutagenicity of 8-OH-dGTP in mutT cells. In this study, 8-OH-dGTP was introduced into wild type and mutT E. coli cells, and mutations of a chromosomal gene were monitored. 8-OH-dGTP induced muta-tions of the rpoB gene, the degree of the mutation induction in the mutT strain being_??_6-fold higher than that in the wild type strain. On the other hand, 2-hydroxy-dATP, which is not a substrate of the MutT protein, increased the mutation to similar degrees in the two strains. These results constitute the first evidence that the MutT protein suppresses mutation by 8-OH-dGTP in vivo.View full abstractDownload PDF (1695K) -
Reconstitution and Replacement of Bacteriochlorophyll α Molecules in Photosynthetic Reaction CentersMasayuki Kobayashi, Atsuo Takaya, Nobuyuki Kanai, Yosuke Ota, Takayuki ...2004 Volume 136 Issue 3 Pages 363-369
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSReaction centers (RCs) of the photosynthetic bacterium Rhodobacter sphaeroides R-26 were reconstituted in liposomes after release of pigments (bacteriochlorophyll α (BChlα) and bacteriopheophytin α (BPheα)) by treatment with acetone. As shown by absorption and circular dichroism spectroscopies, the reconstituted RCs had the same arrangement of pigments as the native RC and exhibited photoactivity of the special pair. The recovery yield of RCs of up to 30% was achieved by addition of 7.8-fold excess of BChlα in the acetone treatment. Furthermore BChlα was partially replaced with Zn-BChlα by addition of the pigments during the acetone treatment. About 30% and 50% of the special pair and accessory pigments can be replaced with Zn-BChlα, respectively. From this rate, an oxidation-reduction potential of 520mV (vs. the normal hydrogen electrode NHE) was derived by the simulation of the exper-imental data, which is 35mV higher than that of the native RC (484mV vs. NHE).View full abstractDownload PDF (838K) -
Da-Huang Chen, Kuan-Tsu Wu, Chien-Jen Hung, Mingli Hsieh, Chuan Li2004 Volume 136 Issue 3 Pages 371-376
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSAdenosine dialdehyde (AdOx) is an indirect methyltransferase inhibitor broadly used in cell culture to accumulate methyl-accepting proteins in hypomethylated states for in vitro protein methylation analyses. In this study we included a translation inhibi-tor, cycloheximide, in the AdOx treatment of HeLa cells. The methyl-accepting proteins disappeared in the double treatment, indicating that they were most likely newly synthesized in the AdOx incubation period. AdOx treatment could also be used in combination with in vivo methylation, another technique frequently used to study protein methylation. AdOx treatment prior to in vivo methylation accumulated methyl-accepting proteins for the labeling reaction. The continued presence of AdOx in the in vivo labeling period decreased the methylation of the majority of in vivo methyl-accepting polypeptides. The level and pattern of the in vivo methylated polypeptides did not change after a 12-h chase, supporting the notion that the meth-ylated polypeptide as well as the methyl groups on the modified polypeptides are stable. On the other hand, methylarginine-specific antibodies detected limited but consistent reduction of the methylarginine-containing proteins in AdOx-treated samples compared to the untreated ones. Thus, AdOx treatment probably only blocked a small fraction of stable protein methylation. Overall, it is likely that base-stable methyla-tion are formed soon after the synthesis of the polypeptide and remain stable after the modification.View full abstractDownload PDF (4862K) -
Hiroko Kojima, Toshimitsu Uede, Toshimasa Uemura2004 Volume 136 Issue 3 Pages 377-386
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSOsteopontin (OPN) is a highly acidic secreted phosphoprotein that binds to cells via an RGD (arginine-glycine-aspartic acid) cell adhesion sequence that recognizes the αvβ3 integrin. OPN may regulate the formation and remodeling of bone. To elucidate the function of OPN in bone tissue, we examined the overexpression of OPN in osteob-lasts in vitro and in vivo using an adenoviral vector carrying an OPN cDNA (Adv-OPN). Rat bone marrow-derived osteoblasts infected with Adv-OPN were examined by Western blotting, immunofluorescence, nodule formation measurements, assay of alkaline phosphatase (ALP) activity, and Northern blotting. The results suggested that not only osteoblast differentiation markers such as osteocalcin and ALP, but nodule formation and ALP activity are markedly enhanced by OPN overexpression in the case of viral infection. On the contrary, when Adv-OPN and uninfected osteoblasts were implanted into subcutaneous sites with a porous ceramic scaffold, the ALP activity and calcium content of the OPN-infected composite were higher than in unin-fected composites, however, the differences were smaller than expected from the in vitro experiments. We speculate that the difference in the result of in vitro and in vivo experiments originates from the inhibitory effect of secreted OPN on the crystal growth of apatite in vivo, which competes with the induced activity of osteoblasts.View full abstractDownload PDF (5328K) -
Kana Shimane, Mitsuo Ogura2004 Volume 136 Issue 3 Pages 387-397
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSThe DegS-DegU two-component system in Bacillus subtilis regulates exoprotease production and competence development. Phosphorylated and unphosphorylated forms of DegU are required for activation of aprE and comK, respectively. Alanine-scanning mutagenesis of the helix-turn-helix region of DegU and in vivo examination of 27 DegU variants revealed five common mutants that showed severe reduction of gene expression of both aprE and comK because of reduced DNA-binding activity. This observation suggested that the DegU-recognized cis-sequences might not be consider-ably changed for either promoter. We identified a DegU-recognized inverted repeat in the comK promoter using various mutant comK-lacZ fusions. Inspection of the aprE promoter sequence revealed a tandem repeat consisting of short AT-rich sequences containing a consensus one, 5'-TAAAT-3', which was found in the downstream half of the inverted repeat involved in comK activation. Oligonucleotide-directed replace-ment of the short AT-rich sequences located in the center of each motif decreased DegU-dependent aprE expression, implying that the repeat is required for the activation of aprE. Based on these results, it was concluded that DegU would function through the inverted repeat in the comK promoter and the tandem repeat in the aprE promoter.View full abstractDownload PDF (3753K) -
Takenori Takizawa, Chizuru Tatematsu, Kimi Watanabe, Kanefusa Kato, Yo ...2004 Volume 136 Issue 3 Pages 399-405
Published: September 01, 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSCalnexin is an endoplasmic reticulum (ER)-resident molecular chaperone that plays an essential role in the correct folding of membrane proteins. We found that calnexin is subjected to partial cleavage in apoptotic mouse cells. Both ER stress-inducing and ER stress-non-inducing apoptotic stimuli caused the cleavage of calnexin, indicating that this event does not always occur downstream of ER stress. The inhibition of caspases that target the amino acid sequence DXXD abrogated calnexin cleavage in apoptotic stimulus-treated cells. In addition, disruption of one of two DXXD sequences located in the cytoplasmic domain caused calnexin to escape cleavage during apoptosis. Furthermore, calnexin was cleaved in vitro by recombinant caspase-3 or caspase-7. Finally, the overexpression of a presumed cleavage product of calnexin partly inhibited apoptosis. These results collectively suggest that caspase-3 or cas-pase-7 cleaves calnexin, whose cleaved product leads to the attenuation of apoptosis.View full abstractDownload PDF (6999K) -
2004 Volume 136 Issue 3 Pages 407
Published: 2004
Released on J-STAGE: June 30, 2008
JOURNAL FREE ACCESSDownload PDF (19K)
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