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61.
Mariko Kato Nahoko Nagasaki-Takeuchi Yuki Ide Rie Tomioka Masayoshi Maeshima 《Plant signaling & behavior》2010,5(7):848-850
In plants, Ca2+, phosphatidylinositol phosphates (PtdInsPs) and inositol phosphates are major components of intracellular signaling. Several kinds of proteins and enzymes, such as calmodulin (CaM), protein kinase, protein phosphatase, and the Ca2+ channel, mediate the signaling. Two new Ca2+-binding proteins were identified from Arabidopsis thaliana and named PCaP1 and PCaP2 [plasma membrane (PM)-associated Ca2+(cation)-binding protein 1 and 2]. PCaP1 has an intrinsically disordered region in the central and C-terminal parts. The PCaP1 gene is expressed in most tissues and the PCaP2 gene is expressed predominantly in root hairs and pollen tubes. We recently demonstrated that these proteins are N-myristoylated, stably anchored in the PM, and are bound with phosphatidylinositol phosphates, especially PtdInsP2s. Here we propose a model for the switching mechanism of Ca2+-signaling mediated by PtdInsPs. Ca2+ forms a complex with CaM (Ca2+-CaM) when there is an increase in the cytosol free Ca2+. The binding of PCaPs with Ca2+-CaM causes PCaPs to release PtdInsPs. Until the release of PtdInsPs, the signaling is kept in the resting state.Key words: calcium signal, calmodulin, inositol phosphate, intrinsically disordered protein, myristoylation, phosphatidylinositol phosphate, plasma membrane 相似文献
62.
We developed a technique for detecting the heat-labile I (LTI) and heat-stable I (STI) genes of enterotoxigenic Escherichia coli (ETEC) using a novel DNA amplification procedure designated Loop-Mediated Isothermal Amplification (LAMP). The detection limit of accelerated LAMP utilizing loop primers was 4 CFU/test for LTI and was 40 CFU/test for STI, which are 10-fold higher than those of conventional PCR assay (detection limit, 40 CFU/test and 400 CFU/test, respectively). No DNA amplification was observed in LT and ST non-producing E. coli or other bacterial strains; thus, high specificity was verified. The specificity of LAMP assay was also confirmed by digestion of LAMP products using restriction enzymes and DNA sequence analysis. In the accelerated LAMP assay, DNA amplification was detected within 35 min, and thus LAMP is superior to conventional PCR in terms of rapidity. It was confirmed that increased concentrations of primers and Bst DNA polymerase could further facilitate the reaction. Furthermore, with the high amplification efficiency of the LAMP assay, amplification can be visually observed by the turbidity caused by magnesium pyrophosphate, a byproduct of the reaction. Detection of LTI and STI in ETEC by LAMP is thus an extremely rapid procedure with high sensitivity and specificity that requires no specialized equipment. This assay is expected to become a valuable tool for rapid diagnosis in ETEC infection. 相似文献
63.
Ca2+-signaling in downstream effectors is supported by many kinds of Ca2+-binding proteins, which function as a signal mediator and a Ca2+-buffering protein. We found in Arabidopsis thaliana a new type of Ca2+-binding protein, CCaP1, which consists of 152 amino acid residues, and binds (45)Ca2+ even in the presence of a high concentration of Mg2+. We found two other proteins with similar motifs, CCaP2 and CCaP3. These three proteins had no organelle localization signal and their green fluorescent protein (GFP) fusions were detected in the cytosol. Real-time PCR and histochemical analysis of promoter-beta-glucuronidase fusions revealed that CCaP1 was predominantly expressed in petioles while CCaP2 was expressed in roots. CCaP3 was hardly expressed. Expression of CCaP1 and CCaP2 was enhanced in darkness and became maximal after 24 h. Immunoblotting revealed petiole-specific accumulation of CCaP1. Expression of CCaP1 and CCaP2 was suppressed by a high concentration of Ca2+ and other metal ions. Deletion of sucrose from the medium markedly increased the mRNA levels of CCaP1 and CCaP2 within 2 h. Gibberellic acid enhanced the expression of CCaP1 and CCaP2 by 5- and 2.5-fold, respectively, after 6 h. CCaP1 and CCaP2 were suppressed in the petiole and the root, respectively, by light and the product of photosynthesis (sucrose) or both. These results suggest that CCaP1 functions as a mediator in response to continuous dark or gibberellic acid. 相似文献
64.
Ohta T Eguchi R Suzuki A Miyakaze S Ayuzawa R Kaji K 《Journal of cellular physiology》2007,211(3):673-681
In order to improve medical treatment of ischemic injury such as myocardial infarction, it is important to elucidate hypoxia-induced changes to endothelial cells. An in vitro blood vessel model, in which HUVECs are stimulated to form a network of capillary-like tubes, was used to analyze hypoxia-induced morphological and biochemical changes. When exposed to hypoxia, the network of capillary tubes broke down into small clusters. This tube breakdown was accompanied by chromatin condensation and cell nuclear fragmentation, morphological markers of apoptosis, and activation of two apoptotic signals, caspase-3 and p38. We investigated what roles caspase cascade and p38 play in hypoxia-induced apoptosis and tube breakdown by using zVAD-fmk and SB203580, specific inhibitors of these two apoptotic signals, respectively. Chromatin condensation and cell nuclear fragmentation and tube breakdown were effectively inhibited by SB203580, but not by zVAD-fmk. SB203580 caused dephosphorylation of p38, which indicates that p38 was autophosphorylated. Inhibition by zVAD-fmk caused slight MW increase in p17 and emergence of p19, which indicates that the inhibitor caused partial processing of caspase-3. Inhibition of p38 suppressed activation of caspase-3 but not vice versa. In addition, these two inhibitors were shown to differentially inhibit cleavage of so-called caspase substrates. SB203580 inhibited cleavage of PARP and lamin A/C, while zVAD-fmk inhibited cleavage of lamin A/C but not that of PARP. Taken together, these results show that p38 is located upstream of caspase cascade and that, although caspase-3 is activated, a p38-regulated caspase-independent pathway is crucial for the execution of hypoxia-induced apoptosis and tube breakdown. 相似文献
65.
Protein kinase C delta regulates the phosphorylation of heat shock protein 27 in human hepatocellular carcinoma 总被引:1,自引:0,他引:1
Takai S Matsushima-Nishiwaki R Tokuda H Yasuda E Toyoda H Kaneoka Y Yamaguchi A Kumada T Kozawa O 《Life sciences》2007,81(7):585-591
We have recently reported that attenuated phosphorylation of heat shock protein (HSP) 27 correlates with tumor progression in patients with hepatocellular carcinoma (HCC). In the present study, we investigated what kind of kinase regulates phosphorylation of HSP27 in human HCC-derived HuH7 cells. 12-O-tetradecanoylphorbol-13-acetate (TPA) and 1-oleoyl-2-acetylglycerol, direct activators of protein kinase C (PKC), markedly strengthened the phosphorylation of HSP27. Bisindorylmaleimide I, an inhibitor of PKC, suppressed the TPA-induced levels of HSP27 phosphorylation in addition to its basal levels. Knock down of PKCdelta suppressed HSP27 phosphorylation, as well as p38 mitogen-activated protein kinase (MAPK) phosphorylation. SB203580, an inhibitor of p38 MAPK, suppressed the TPA-induced HSP27 phosphorylation. Our results strongly suggest that activation of PKCdelta regulates the phosphorylation of HSP27 via p38 MAPK in human HCC. 相似文献
66.
Vascular plants have evolved shoot apical meristems (SAMs), whose structures differ among plant groups. To clarify the evolutionary course of the different structural types of SAMs, we compared plasmodesmatal networks in the SAMs for 17 families and 24 species of angiosperms, gymnosperms, and pteridophytes, using transmission electron microscopy (TEM). The plasmodesmata (PD) in almost all cell walls in median longitudinal sections of SAMs were counted, and the PD density per unit area was calculated for each cell wall. Angiosperm and gymnosperm SAMs have low densities, with no difference between stratified (tunica-corpus) and unstratified structures. SAMs of ferns, including Psilotum and Equisetum, have average densities that are more than three times higher than those of seed plants. Interestingly, microphyllous lycopods have both the fern and seed-plant types of PD networks; Selaginellaceae SAMs with single apical cells have high PD densities, while SAMs of Lycopodiaceae and Isoetaceae with plural initial cells have low PD densities, equivalent to those of seed plants. In summary, PD networks are strongly correlated to SAM organizations-SAMs with single and plural initial cells have the fern and seed-plant types of PD, respectively. The two SAM organizations may have evolved separately in lycophytes and euphyllophytes and may be associated with gain or loss of the ability to form secondary PD. 相似文献
67.
Okada Y Imendra KG Miyazaki T Hotokezaka H Fujiyama R Zeredo JL Toda K 《Cellular and molecular neurobiology》2007,27(6):771-781
The effect of calcium-sensing receptor (CaR) agonists on frog gustatory responses was studied using glossopharyngeal nerve
recording and whole-cell patch-clamp recording of isolated taste disc cells. Calcimimetic NPS R-467 dissolved in normal saline
solution elicited a large transient response in the nerve. The less active enantiomer of the compound, NPS S-467 induced only
a small neural response. The EC50 for NPS R-467 was about 20 μM. Cross-adaptation experiments were performed to examine the effect of 30 μM NPS R-467 and 100 μM
quinine on the gustatory neural response. The magnitude of the R-467-induced response after adaptation to quinine was approximately
equal to that after adaptation to normal saline solution, indicating that the receptor site for NPS R-467 is different from
the site for quinine. NPS R-467 (100 μM) also induced an inward current accompanied with conductance increase and large depolarization
in two (13%) of 15 rod cells, and a sustained decrease in outward current and small depolarization in six (40%) other rod
cells. NPS S-467 (100 μM) induced a sustained decrease in outward current and depolarization in five (50%) of 10 rod cells.
Another calcimimetic cinacalcet (100 μM) induced an inward current accompanied with conductance increase in three (27%) of
11 rod cells. The results suggest that NPS R-467 induces neural responses through cell responses unrelated to a resting K+ conductance decrease. 相似文献
68.
Hayakawa T Kanagawa R Kotani Y Kimura M Yamagiwa M Yamane Y Takebe S Sakai H 《Current microbiology》2007,55(4):278-283
A novel crystal protein that exhibited potent cytotoxicity against human leukemic T-cells was cloned from the Bacillus thuringiensis TK-E6 strain. The protein, designated as parasporin-2Ab (PS2Ab), was a polypeptide of 304 amino acid residues with a predicted
molecular weight of 33,017. The deduced amino acid sequence of PS2Ab showed significant homology (84% identitiy) to parasporin-2Aa
(PS2Aa) from the B. thuringiensis A1547 strain. Upon processing of PS2Ab with proteinase K, the active form of 29 kDa was produced. The activated PS2Ab showed
potent cytotoxicity against MOLT-4 and Jurkat cells and the EC50 values were estimated as 0.545 and 0.745 ng/mL, respectively. The cytotoxicity of PS2Ab was significantly higher than that
of PS2Aa reported elsewhere. Although both cytotoxins were structurally related, it was thought that the minor differences
found were responsible for the different cytotoxicities of PS2Ab and PS2Aa. 相似文献
69.
Xiao-Jie Li Rie Uenishi Saiki Hase Huanan Liao Tee Kok Keng Shigeru Kusagawa Yutaka Takebe 《Virologica Sinica》2007,22(6):426-433
The Asia-Pacific region is a home to 60% of the population in the world and to approximately one quarter of people with HIV/AIDS. Close to a million of people has been infected and a half million people died of AIDS annually in Asia,becoming the second largest epicenter of global AIDS epidemic. Molecular epidemiology has been useful tool to track a course of HIV spread. In-depth knowledge from the studies on molecular epidemiology elucidates the dynamics of HIV spread and the interrelationship of epidemics in the different regions in Asia. 相似文献
70.
Kuratani M Yoshikawa Y Bessho Y Higashijima K Ishii T Shibata R Takahashi S Yutani K Yokoyama S 《Structure (London, England : 1993)》2007,15(12):1642-1653
In the bacterial genetic-code system, the codon AUA is decoded as isoleucine by tRNA(Ile)(2) with the lysidine residue at the wobble position. Lysidine is derived from cytidine, with ATP and L-lysine, by tRNA(Ile) lysidine synthetase (TilS), which is an N-type ATP pyrophosphatase. In this study, we determined the crystal structure of Aquifex aeolicus TilS, complexed with ATP, Mg2+, and L-lysine, at 2.5 A resolution. The presence of the TilS-specific subdomain causes the active site to have two separate gateways, a large hole and a narrow tunnel on the opposite side. ATP is bound inside the hole, and L-lysine is bound at the entrance of the tunnel. The conserved Asp36 in the PP-motif coordinates Mg2+. In these initial binding modes, the ATP, Mg2+, and L-lysine are held far apart from each other, but they seem to be brought together for the reaction upon cytidine binding, with putative structural changes of the complex. 相似文献