Neutrophil activation and collagen metabolism in lung cancer.
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Adenocarcinoma↗
Biomedical subjects
M Yoshida
Publications and source records attributed to M Yoshida.
F1-ATPase alpha-subunit made up from two fragments (1-395, 396-503) is stabilized by ATP and complexes containing it obey altered kinetics.
Inferred from the crystal structure of mitochondrial F1-ATPase (Abrahams, J.P. et al. (1994) Nature 370, 621-628), the proteinase-sensitive region around Phe-395 of thermophilic F1-ATPase alpha-subunit corresponds to the loop which connects main part of the carboxyl-terminal helical bundle domain with the ATP binding domain. This loop is in contact with the gamma- and adjacent beta-subunits. Two polypeptides corresponding to the sequence 1-395 and 396-503 of the alpha-subunit were expressed in Escherichia coli cells and they were copurified as an apparently functional alpha-subunit (alpha(395/396)) made up of two polypeptides. The isolated alpha(395/396) was stabilized by ATP-Mg, but not by ADP-Mg, although it bound both ATP-Mg and ADP-Mg with similar affinities (Kd, 11 microM and 14 microM, respectively). The alpha(395/396) was reconstitutable into alpha(395/396)3 beta 3 and alpha(395/396)3 beta 3 gamma complexes. Different from the intact the ATP-Mg-induced dissociation into alpha 1 beta 1 heterodimers. ATP hydrolysis by the alpha(395/396)3 beta 3 gamma complex underwent a slow initial phase, whereas the intact alpha 3 beta 3 gamma complex exhibited an accelerated initial phase. Steady-state ATPase activity at various ATP concentrations showed negative cooperativity for the intact alpha 3 beta 3 gamma complex but apparently positive cooperativity for the alpha(395/396)3 beta 3 gamma complex. The ATPase activities at a saturating ATP concentration of the complexes containing the alpha(395/396) were 180% of those containing intact alpha-subunits. These results indicate that a loop around Phe-395 is involved in intersubunit interaction in F1-ATPase.
Adenosine Diphosphate↗
Neuronal mechanism underlying dystonia induced by bicuculline injection into the putamen of the cat.
Microinjection of bicuculline (Bic) into the unilateral putamen (Put) of the cat resulted in periodic occurrence of contralateral movement of the head (dystonia). Corresponding to this dystonic movement, repetitive spikes of the field potential occurring in the Put of the injected side and strong inhibition of the extracellularly recorded unit discharges in the neurons of the substantia nigra pars reticulata (SNr) of the same side were observed.
Animals↗
Stimulation of transcription accompanying relaxation of chromatin structure in cells overexpressing high mobility group 1 protein.
We developed murine C-127 cell lines that stationarily overexpress high mobility group (HMG) proteins 1 and 2 by transfecting them with the bovine papilloma virus plasmid carrying their respective cDNA sequences. Using these cell lines, we examined the effects of these HMG proteins on the modulation of chromatin structure that accompanied transcription. The levels of HMG1 mRNA and protein in cells overexpressing HMG1 protein were enhanced about 7- and 3-fold, respectively, in comparison with control cells, whereas those in cells overexpressing HMG2 protein were enhanced about 17- and 9-fold. The expression of reporter genes transfected into the cells was enhanced approximately 2-fold in cells overexpressing HMG1, but not HMG2, in comparison with those in control cells, irrespective of the sources of the genes and promoters. The minichromosome derived from the reporter plasmid in cells overexpressing HMG1 protein was more susceptible to micrococcal nuclease digestion than those in cells overexpressing HMG2 protein and control cells. The enhanced accessibility to micrococcal nuclease was not restricted to the expressing gene and promoter but involved the entire minichromosome, suggesting that the enhancement of gene expression resulted from changes in the condensation of the entire minichromosomal region by HMG1 protein. Minichromosomes in cells overexpressing HMG contained enhanced amounts of the respective HMG proteins and simultaneously reduced amounts of histone H1s. These results suggest that HMG1 and -2 proteins have different functions in the modulation of chromatin structure, and that HMG1 protein may sustain the structure of the respective gene to ensure that its activity as a template is expressed fully. These observations on the modulation of chromatin structure accompanying gene transcription in cells overexpressing HMG protein may provide important information on the function of these proteins.
Animals↗
Equatorial split of holo-chaperonin from Thermus thermophilus by ATP and K+.
Holo-chaperonin molecule from Thermus thermophilus is a bullet-shaped particle whose cylinder part and round top are composed of two stacked rings of the cpn60 heptamer and a single ring of the cpn10 heptamer, respectively. We found that it splits at the plane between two cpn60 rings into two halves under physiological conditions, that is, in the presence of ATP (but not AMP-PNP, ADP) + K+ (but not Na+) at 60 degrees C. This equatorial split could be functionally important although it has not been considered in any current mechanistic model of chaperonin functioning.
Adenosine Triphosphate↗
Human T-cell leukemia virus type-1 (HTLV-1) Tax is expressed at the same level in infected cells of HTLV-1-associated myelopathy or tropical spastic paraparesis patients as in asymptomatic carriers but at a lower level in adult T-cell leukemia cells.
Patients with human T-cell leukemia virus type-1 (HTLV-1)-associated myelopathy (HAM/TSP) generally harbor a much greater population of HTLV-1-infected T cells in peripheral blood mononuclear cells (PBMCs) than asymptomatic carriers. These cells are not malignant but frequently proliferate clonally. To investigate the possibility that higher expression of the viral activator Tax induces preferential proliferation of infected nonmalignant T cells in HAM/TSP patients, the expression of Tax mRNA in fresh PBMCs was analyzed by reverse transcriptase-mediated polymerase chain reaction. Total amount of Tax mRNA was higher in HAM/TSP patients than in carriers, but the expression level was almost the same as that in asymptomatic carriers when compared per infected cell. The expression levels in adult T-cell leukemia patients were significantly lower than those in HAM/TSP patients and asymptomatic carriers. These results indicate that tax gene is not expressed at a continuously high level in HAM/TSP patients who carry a high population of infected T cells, even in those with clonally proliferated infected T cells.
Base Sequence↗
Tax protein of HTLV-1 destabilizes the complexes of NF-kappa B and I kappa B-alpha and induces nuclear translocation of NF-kappa B for transcriptional activation.
Tax protein of HTLV-1 stimulates transcription of specific cellular genes through the NF-kappa B binding site. We previously showed that Tax binds to the ankyrin motifs of I kappa B-gamma, and dissociates the I kappa B-gamma/NF-kappa B complexes, resulting in nuclear translocation of NF-kappa B proteins. We herein report the effects of Tax on I kappa B-alpha, another member of I kappa B family proteins expressed in human T cells. Tax binds to I kappa B-alpha and partially dissociates the NF-kappa B/I kappa B-alpha complex in vitro. In Tax-expressing cells, Tax/I kappa B-alpha complex was detected only at low level, but NF-kappa B/I kappa B-alpha complex was mostly dissociated and NF-kappa B was translocated into the nucleus. Furthermore, Tax induced reduction of I kappa B-alpha protein. I kappa B-alpha protein was stabilized by NF-kappa B protein through forming NF-kappa B/I kappa B-alpha complex, but Tax expression cancelled this stabilization effect on I kappa B-alpha. These findings suggest that Tax induces dissociation of and/or inhibits the formation of NF-kappa B/I kappa B-alpha complexes, resulting in release of NF-kappa B and destabilization of I kappa B-alpha. Consistently, transcription of NF-kappa B directed gene was activated. These Tax effects counteract the negative feedback control of NF-kappa B by I kappa B-alpha and contribute to constitutive activation of NF-kappa B in Tax-transfected and HTLV-1-infected cells.
Biological Transport↗
Radicicol, a protein tyrosine kinase inhibitor, suppresses the expression of mitogen-inducible cyclooxygenase in macrophages stimulated with lipopolysaccharide and in experimental glomerulonephritis.
Two isoforms of cyclooxygenase (COX) have been identified in eukaryotic cells: a constitutively expressed COX-1 and mitogen-inducible COX-2, which is selectively expressed in response to various inflammatory stimuli. Thus, COX-2 instead of COX-1 is implicated to produce prostanoids mediating inflammatory responses. Major efforts have been focused on identifying nonsteroidal anti-inflammatory drugs (NSAIDS) which can selectively inhibit the enzyme activity of COX-2. Such NSAIDS would be more desirable anti-inflammatory agents in comparison to NSAIDS which inhibit both COX-1 and COX-2. Other than glucocorticoids, pharmacological agents which can selectively suppress the expression of COX-2 without affecting that of COX-1 have not been identified. We report here that radicicol, a fungal antibiotic, is a potent protein tyrosine kinase inhibitor, and that it inhibits the expression of COX-2 without affecting COX-1 expression in lipopolysaccharide (LPS)-stimulated macrophages with the IC50 value of 27 nM. Radicicol inhibited tyrosine phosphorylation of p53/56lyn, a Src family tyrosine kinase and one of the major tyrosine-phosphorylated proteins in LPS-stimulated macrophages. Radicicol also inhibited COX-2 expression in vivo in glomeruli of rats with experimental glomerulonephritis induced by the anti-glomerular basement membrane antibodies, in which COX-2 expression is known to be enhanced. The enzyme activity of COX-1 or COX-2 was not affected by radicicol in macrophages. Radiciciol also suppressed the COX-2 expression induced by IL-1 beta in rat smooth muscle cells. Other protein tyrosine kinase inhibitors suppressed the LPS-induced COX-2 expression in macrophages but at much higher concentrations than needed for radicicol. Radicicol did not inhibit the COX-2 expression induced by phorbol 12-myristate 13-acetate in macrophages. These results suggest that the activation of tyrosine-specific protein kinases is the proximal obligatory step in the LPS-induced signal transduction pathway leading to the induction of COX-2 expression in macrophages. The magnitude of the inhibition of COX-2 protein synthesis by radicicol was much greater than that of the steady state levels of COX-2 mRNA. These results suggest that radicicol inhibits COX-2 expression mainly at post-transcriptional steps.
Animals↗
Urinary excretion of N-acetyl-beta-D-glucosaminidase during normal pregnancy.
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Acetylglucosaminidase↗
Existence of a transcription factor for the human HMG2 gene positively related to the level of HMG2 mRNA in the cells.
A functional gene encoding high mobility group 2 (HMG2) protein, which is an abundant eukaryotic DNA-binding protein, has been isolated. The expression of the HMG2 gene is enhanced in exponentially growing cells and in cells transformed with various viral genes and oncogenes. We attempted to identify and characterize the HMG2 gene structure and transcription factor(s) participating in the expression of the gene. Chloramphenicol acetyltransferase assays to characterize the 5'-flanking region of the human HMG2 gene revealed that the nucleotide sequences in two regions are necessary for expression of the HMG2 gene: one (-85 to +44 region) as a core promoter, and the other (-621 to -493 region) as a cis regulatory element(s). Electrophoresis mobility shift assay with a DNA fragment containing the cis regulatory element and a crude nuclear extract from HeLa cells gave several complexes. Chemical footprint and competition assays indicated that the component giving one of the major complexes recognizes a nucleotide sequence of -499 to -486 in the cis regulatory element. Chloramphenicol acetyltransferase assay indicated that the component giving the major complex is required for the effective transcription of the HMG2 gene. The binding component named HMG2TF (HMG2 transcription factor) contained a protein in apparent molecular size of 85,000, as determined by a UV cross-linking experiment. The amount of HMG2TF in the growing cells and transformed cells increased in positive relationship to the level of expression of HMG2 mRNA in the cells. These results suggest that HMG2 gene expression may be regulated by the relative amount of this transcription factor.
Base Sequence↗
Neurotrophic effects of conditioned media of astrocytes isolated from different brain regions on hippocampal and cortical neurons.
The present study was designed to reveal whether astroglial cells from different brain regions produce diffusible factors that differentially support the survival of neurons and the establishment of neuronal morphology. For this purpose, astrocyte conditioned media (ACM) were prepared by conditioning chemically-defined medium with type I astrocyte culture dissociated from cerebral cortex, hippocampus and hypothalamus. Hippocampal and cortical neurons were cultured in ACM or in non-conditioned medium. ACM derived from three brain regions all supported the survival of hippocampal and cortical neurons better than non-conditioned control medium. Of these, hypothalamic ACM was the most effective in supporting the survival of cortical neurons. The ACM also potentiated the elongation of the longest neurite of hippocampal and cortical neurons. However, there were no significant differences in the promoting effects on neurite elongation among ACM from three brain regions.
Animals↗
Chaperonin releases the substrate protein in a form with tendency to aggregate and ability to rebind to chaperonin.
To know whether the protein released from chaperonin GroEL/ES is in a form committed to the native state or still an aggregatable non-native one, two experiments were carried out. Dilution of the [GroEL-substrate protein] binary complex prior to ATP addition significantly improved the yield of folding, suggesting that the released protein has a tendency to aggregate. When N-ethylmaleimide treated GroEL, which can form the binary complex but not release the bound protein, was added to the binary complex prior to ATP addition, productive folding was severely inhibited, indicating that the protein released from GroEL/ES can bind to N-ethylmaleimide treated chaperonin. These data favor the 'reservoir' or 'reversion' model, in which GroEL/ES acts as a buffer of folding intermediate or mediates reversion of a misfolded protein to a less folded primitive form, rather than the 'marsupium' model in which folding of the substrate protein proceeds in chaperonin.
Adenosine Triphosphate↗
A common topology of proteins catalyzing ATP-triggered reactions.
A protein fold, six parallel beta strands surrounding the central alpha helix, is likely to be a common structure in protein families known to have a typical set of nucleotide binding consensus sequence motifs A and B and to catalyze ATP-triggered reactions. According to this ATP-triggered protein fold, the conserved Glu (or Asp), which acts as a general base to activate a water molecule for an in-line attack of the gamma-phosphate, is at the exit of the second beta strand. The fifth beta strand may be involved in propagation of conformational change triggered by ATP hydrolysis.
Adenosine Triphosphate↗
HTLV-1 Tax enhances NF-kappa B2 expression and binds to the products p52 and p100, but does not suppress the inhibitory function of p100.
Tax protein of HTLV-1 triggers transcriptional activation through enhancers, NF-kappa B binding site, 21-bp enhancer, and serum response element. Previously, we demonstrated binding of Tax to transcription factors NF-kappa B1 p105 and p50. Here, we report that Tax enhances expression of NF-kappa B2 at the mRNA level and proteins; the effect was more apparent on the p52 expression than on its precursor p100, suggesting post-translational regulation. Consistent with these observations, HTLV-1-infected T-cell lines expressed higher levels of p52. Tax binds to the protein products p52 and p100 which inhibits NF-kappa B proteins forming cytoplasmic complexes; the binding to p100 was preferential over NF-kappa B1 p105. However, Tax did not induce efficient dissociation of the cytoplasmic complexes p100/c-Rel or p100/p65, and thus did not induce nuclear translocation of c-Rel or p65. This was in sharp contrast to the previous observation that Tax dissociated the p105/c-Rel and I kappa B-gamma/p65 complexes. These results indicate that HTLV-1 Tax interacts with NF-kappa B2 p100 and p52 and upregulate the NF-kappa B function, but their contribution to Tax-mediated transcriptional regulation differs from those of NF-kappa B1.
Base Sequence↗
Plasma (1-->3)-beta-D-glucan measurement in diagnosis of invasive deep mycosis and fungal febrile episodes.
(1-->3)-beta-D-glucan is a characteristic fungal cell-wall constituent. To assess the clinical usefulness of this glucan in screening for invasive fungal infection or fungal febrile episodes, we measured the plasma concentration at the time of routine blood culture in 202 febrile episodes by means of factor G, a horseshoe-crab coagulation enzyme that is extremely sensitive to this polysaccharide. With a plasma cut-off value of 20 pg/mL, 37 of 41 episodes of definite fungal infections (confirmed at necropsy or by microbiology) had positive results (sensitivity 90%). All of 59 episodes of non-fungal infections, tumour fever, or collagen diseases had concentrations below the cut-off value (specificity 100%). Of 102 episodes of fever of unknown origin, 26 had plasma glucan concentrations of more than 20 pg/mL. With those 102 cases taken as non-fungal infections, the positive predictive value of the test was estimated as 59% (37/63), the negative predictive value as 97% (135/139), and the efficiency as 85% (172/202). The positive predictive value should improve if there were a sensitive gold standard that could discriminate fungal from non-fungal infections. Causative fungi included candida, aspergillus, cryptococcus, and trichosporon. Determination of plasma (1-->3)-beta-D-glucan with factor G is a highly sensitive and specific test for invasive deep mycosis and fungal febrile episodes, and will substantially benefit immunocompromised patients.
Adolescent↗
HLA-DRB1 frequencies of non-Austronesian-speaking Gidra in south New Guinea and their genetic affinities with Oceanian populations.
DNA typing of HLA-DRB1 genes was conducted for 192 samples from the Gidra who speak one of the non-Austronesian languages and inhabit the southern lowlands of New Guinea. Comparison of the allele frequencies with those of eight other Oceanian populations reveals that the Gidra are genetically closest to the non-Austronesian-speaking Goroka and, next, to the Aboriginal Australian groups, but are remote from the Austronesian-speaking groups in mainland New Guinea and the rest of Melanesia. This finding clearly supports the hypothesis that non-Austronesians and Aboriginal Australians are descendants of the first-stage migrants to Oceania.
Adolescent↗
Trichostatin A and trapoxin: novel chemical probes for the role of histone acetylation in chromatin structure and function.
Reversible acetylation at the epsilon-amino group of lysines located at the conserved domain of core histones is supposed to play an important role in the regulation of chromatin structure and its transcriptional activity. One promising strategy for analyzing the precise function of histone acetylation is to block the activities of acetylating or deacetylating enzymes by specific inhibitors. Recently, two microbial metabolites, trichostatin A and trapoxin, were found to be potent inhibitors of histone deacetylases. Trichostatin A reversibly inhibits the mammalian histone deacetylase, whereas trapoxin causes inhibition through irreversible binding to the enzyme. The histone deacetylase from a trichostatin A-resistant cell line is resistant to trichostatin A, indicating that the enzyme is the primary target. Both of the agents induce a variety of biological responses of cells such as induction of differentiation and cell cycle arrest. Trichostatin A and trapoxin are useful in analyzing the role of histone acetylation in chromatin structure and function as well as in determining the genes whose activities are regulated by histone acetylation.
Acetylation↗
Correlation between tumor volume doubling time and histologic findings in gastric smooth muscle tumors: clinical implications of tumor volume doubling time.
To assess the clinical implications of the tumor volume doubling time of gastric smooth muscle tumors based on a comparison with the histologic findings, seven tumors (four leiomyomas and three leiomyosarcomas) were followed up by consecutive upper gastrointestinal studies between March 1985 and December 1993. The patients were four men and three women with an average age of 58 years (range: 50-71 years). The observation period ranged from 6 to 51 months, with an average of 35 months. All tumors were surgically resected and the histologic diagnosis was confirmed. The following microscopic features were evaluated: 1) mitotic rate, 2) nuclear atypia, and 3) cellularity. Each tumor was also evaluated for the presence or absence of necrosis, hemorrhage, and degeneration. The doubling time ranged from 5 to 27 months with a mean of 16 months. There was a strong negative correlation between the mitotic rate and the doubling time (r = -0.935, P = 0.0019). The doubling time was also significantly related to nuclear atypia, but the number of tumors studied was so small that its reliability was questionable. The doubling time was not related to any other histologic findings. This study shows that the doubling time is useful for estimating the malignant potential of gastric smooth muscle tumors, and that tumors with a doubling time of 16 months or less should be considered as malignant.
Aged↗