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Biomedical subjects

G Mamiya

Publications and source records attributed to G Mamiya.

At least 19 recordsLinked to original sources
PubMed2000

Detection of kinases that phosphorylate 14-3-3 binding sites of Raf-1 using in situ gel kinase assay.

Raf-1 is a serine/threonine protein kinase that plays a critical role in mitogenic signal transduction. Raf-1 activation requires 14-3-3 binding to Raf-1 as an essential step. This binding is regulated through phosphorylation of Ser259 and Ser621 of Raf-1, each constituting part of the consensus motif for the binding of Raf-1 to 14-3-3. However, Raf-1 kinase kinase(s) that phosphorylates these sites remains unknown. In this report, we detected Raf-1 kinase kinase activity using recombinant glutathione-S-transferase-Raf-1 fusion proteins as substrate of in situ gel kinase assay. Ser259 was phosphorylated by a kinase with a molecular weight of 90 kDa, which was suggested to be Rsk judging from the molecular size, the time course of activation after EGF stimulation and the elution pattern from an anion-exchange column. The Raf-1 fragment containing Ser621 was phosphorylated by kinases with molecular weights of 85, 60, 50 and 48 kDa but not by the kinase that phosphorylates Ser259. These results suggest that although Ser259 and Ser621 lie in the same amino acid sequence motif for 14-3-3 binding, these two regulatory sites for this binding are phosphorylated by different protein kinases.

Animals↗
PubMed1997

The mouse extracellular signal-regulated kinase 2 gene. Gene structure and characterization of the promoter.

ERK2 (extracellular-signal regulated kinase 2, also known as p42 mitogen-activated protein kinase) is an integral member of the mitogen-activated protein kinase cascade that is crucial for many cellular events such as proliferation and differentiation. Here, we determined the genomic organization of the Erk2 gene and characterized its promoter. The Erk2 gene spans over 60 kilobases, and the coding region is split into eight exons. In the coding region, exon-intron organization was exactly conserved between the two mouse genes for ERK2 and ERK1 except one junction shifted by one nucleotide. Primer extension and S1 nuclease analyses identified two major transcription start sites located at -219 and -223 relative to the translation start site. The 5'-flanking sequence lacked TATA box but contained a CCAAT box located approximately 60 base pairs upstream of transcription start sites. Sequencing of the 5'-flanking region also revealed potential cis-acting elements for multiple transcriptional regulatory factors including Sp1, zif268, Ets, CREB, and PuF sites. The promoter activity of the 5'-flanking region was examined using chloramphenicol acetyltransferase as a reporter gene. Transient transfection experiments using Chinese hamster ovary cells defined a maximal promoter activity in a 371-base pair region immediately upstream of the translation start site. Furthermore, we demonstrated, using mouse P19 embryonal carcinoma cells, that this 371-base pair sequence is likely to be sufficient to confer the transcriptional activation of the ERK2 promoter during the retinoic acid-induced differentiation of P19 cells.

Amino Acid Sequence↗
PubMed1995

Potentiating effects of methamphetamine on the hyperactivity of microencephalic rats treated prenatally with methylazoxymethanol: possible implication of hyperdopaminergia.

Microencephalic rats induced by methylazoxymethanol (MAM) were observed to have notable hyperactivity compared to control rats, as measured by several behavioral parameters in an automated field apparatus. Acute injection of the stimulant drug, methamphetamine (MAP), produced an increase in the incidence of locomotion and rearing in control rats, and this stimulatory effect of MAP on motor activity was markedly potentiated in MAM rats. Chronic MAP treatment did not change D1 or D2 dopamine receptor densities in either control or MAM rats. From these results, it was suggested that augmented dopaminergic functions may contribute to the hyperactivity seen in MAM rats.

Alkylating Agents↗
PubMed1994

Immunohistochemical analysis of nm23/NDP kinase expression in human lung adenocarcinoma: association with tumor progression in Clara cell type.

Levels of nm23-H1/nucleoside diphosphate kinase (NDP kinase) expression have been reported to correlate inversely with metastatic potential in some tumors but not in others. Whether or not nm23 gene product is associated with metastatic potential in lung cancer is not clear as yet. We therefore immunohistochemically examined the expression of nm23 gene products in primary lung adenocarcinomas according to cytologic subtypes in order to clarify the association of its expression with the clinical features of the disease. Seventy-two (64.9%) of the 111 lung adenocarcinomas were positive for nm23 protein. In lung adenocarcinoma of Clara cell type, high levels of nm23 expression were associated with advanced pathologic stage, positive lymph node status, and poorer prognosis (P < 0.05). However, no correlation with clinical outcome was observed in other cell types. Our data suggest that higher levels of nm23 expression are associated with tumor progression in lung adenocarcinoma of Clara cell type.

Adenocarcinoma↗
PubMed1993

Immunohistochemical analysis of cathepsin B expression in human lung adenocarcinoma: the role in cancer progression.

Production of cathepsin B by tumor cells has been linked to metastatic potential in several experimental models. Sections of 95 primary lung adenocarcinomas were examined for expression of cathepsin B using a standard avidin-biotin immunohistochemical technique. Staining for cathepsin B was observed in 22.1% of all cases and 28.0% of those of the Clara cell type. In Clara cell adenocarcinomas, cathepsin B expression correlated with positive lymph node status, presence of distant metastases, and poor prognosis (P < 0.05). However, no correlation with clinical outcome was observed in other cell types. Our data suggest that cathepsin B may be involved in invasion and metastasis in Clara cell lung adenocarcinoma.

Adenocarcinoma↗
PubMed1993

Effects of long-term treatment of haloperidol on D2 dopamine receptors in various areas of rat brain.

Effects of long-term treatment of haloperidol on D2 dopamine receptors in various brain areas were examined. Haloperidol treatment was found to increase the number of binding sites of D2 dopamine receptors in striatum, olfactory tubercle, hypothalamus and anterior pituitary, while Bmax of D2 receptors in hippocampus was decreased by haloperidol treatment. Therefore it is suggested that D2 dopamine receptors in hippocampus and other brain areas are regulated differently.

Animals↗
PubMed1992

cDNAs and deduced amino acid sequences of subunits in the binding component of mouse bactericidal factor, Ra-reactive factor: similarity to mannose-binding proteins.

The complement-dependent bactericidal factor, Ra-reactive factor, binds specifically to Ra polysaccharide, which is common to some strains of Gram-negative enterobacteria, and its is a complex of proteins composed of a polysaccharide-binding component and a component that is presumably responsible for the complement activation. The former component consists of two different 28-kDa polypeptides, P28a and P28b. We determined the partial amino acid sequences of P28a and P28b, and the results indicated that these polypeptides were similar to two species of mannose-binding protein, MBP-C and MBP-A (alternative names, liver and serum mannan-binding proteins, respectively), which have been isolated from rat liver and/or serum [Drickamer, K., Dordal, M. S., & Reynolds, L. (1986) J. Biol. Chem. 261, 6878-6887; Oka, S., Itoh, N., Kawasaki, T., & Yamashina, I. (1987) J. Biochem. 101, 135-144]. Thus, we cloned the respective cDNAs, using as probes synthetic oligonucleotides for which the sequences had been deduced from the amino acid sequences of P28a and P28b and of rat MBP cDNAs. The primary structures of P28a and P28b deduced from the cloned cDNAs are homologous to one another. They have three domains, a short NH2-terminal domain, a collagen-like domain, and a domain homologous to regions of some carbohydrate-binding proteins, as has been reported for rat MBPs. Southern and Northern blotting analyses using these cDNAs indicated that the P28a and P28b polypeptides are the products of two unique mouse genes which are expressed in hepatic cells.

Amino Acid Sequence↗
PubMed1992

Effects of foetal treatment with methylazoxymethanol on noradrenergic synapses in rat cerebral cortex.

Methylazoxymethanol (MAM)-induced cerebral hypoplasia resulted in a significant increase in densities of norepinephrine uptake sites in cerebral cortex, suggesting that norepinephrinergic axon terminals were compressed in the smaller brain volumes. The density of beta-adrenergic receptors in MAM-lesioned cerebral cortex was decreased probably due to down-regulation, while there were no changes in the proportions and affinities of agonist high-affinity sites and agonist low-affinity sites in the desensitized beta-receptors.

Alkylating Agents↗
PubMed1991

Purification and partial sequencing of inhibitory factor on renal membrane adenylate cyclase in pancreatic cancer extract: identity with histones H1b or H1d.

Inhibitory activity on renal membrane adenylate cyclase (AC) has previously been found in the extract of a pancreatic cancer associated with humoral hypercalcemia of malignancy (HHM). AC inhibitor was purified employing inhibition of AC activity of renal membrane stimulated by forskolin as its index. N-terminal 9 residues and a digested fragment of purified protein (14 residues) were completely consistent with that of histones H1b and H1d. Not only histone H1 but also histones H2A, H2B and H3 from calf thymus inhibited AC activity. These results indicate that the AC inhibitor in the pancreatic cancer extract is histone H1b or H1d and histones H2A, H2B and H3 also have an AC inhibitory activity.

Adenylyl Cyclases↗
PubMed1991

Organ-specific occurrence and expression of the isoforms of nonspecific lipid transfer protein in castor bean seedlings, and molecular cloning of a full-length cDNA for a cotyledon-specific isoform.

Four kinds of nonspecific lipid transfer proteins (nsLTP) were purified from different organs of castor bean (Ricinus communis L.) seedlings. Amino acid compositions and amino-terminal sequences of the four nsLTPs were determined and compared with those of castor bean isoforms, nsLTP-A, -B, and -C, previously reported [Takishima et al. (1986) Biochim. Biophys. Acta 870, 248-255; Takishima et al. (1988) Eur. J. Biochem. 177, 241-249]. Two isoforms from the cotyledons were identified as nsLTP-A and -C, one isoform from the endosperms as nsLTP-B, and the other was a new isoform from the axes. This new isoform was named nsLTP-D and its amino acid sequence was determined. These results demonstrated organ-specific occurrence of the nsLTP isoforms in castor bean seedlings. The isoforms nsLTP-A, -B, -C, and -D showed similar transfer activity not only for phosphatidylcholine and phosphatidylethanolamine but also for monogalactosyldiacylglycerol, although the homology among their amino acid sequences ranged from 70 to 30%. Two cDNA clones (pnsLTP-C and pnsLTP-D) for nsLTPs of castor bean seedlings were isolated and sequenced. pnsLTP-C was the cDNA clone for nsLTP-C expressed in the cotyledons, and pnsLTP-D was that for nsLTP-D in the axis. A coupled in vitro transcription-translation analysis of both cDNA clones revealed that pnsLTP-C encodes the full-length of nsLTP-C precursor (pro-nsLTP-C), while pnsLTP-D encodes a part of nsLTP-D precursor. PronsLTP-C contained a 24-amino acid pre-sequence preceding the mature nsLTP-C (92 amino acids).(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1990

Increased uptake sites for serotonin and dopamine with decreased S2 serotonin receptors in microencephalic rat brain.

Methylazoxymethanol (MAM)-induced cerebral hypoplasia resulted in a significant increase in densities of both serotonin uptake sites in frontal cortex and dopamine uptake sites in striatum, suggesting serotonergic and dopaminergic axons terminals were compressed in the smaller brain volumes. The density of S2 serotonin receptors in MAM-lesioned frontal cortex was decreased probably due to down-regulation, while densities of D1 and D2 dopamine receptors in striatum were identical between MAM-lesioned rats and control rats.

Abnormalities, Drug-Induced↗
PubMed1989

Dopamine D1 and D2 receptors in spontaneously hypertensive rat brain striatum.

Since it has been reported that dopamine D2 receptors are elevated in the brain striatum of spontaneously hypertensive (SH) rats, and since both D1 and D2 receptors may interact with one another, we measured the densities of both these receptors in SH rat striatum, as well as those in the normotensive Wistar-Kyoto rat striatum. The D1 receptor density in both strains was virtually the same, 72.9 +/- 2.2 and 71.3 +/- 3.2 pmol/g, respectively (mean +/- SD). The D2 receptor densities were also almost identical, 16.3 +/- 0.6 and 16.8 +/- 1.0 pmol/g, respectively (mean +/- SD). Thus, these data do not support the concept of a dopamine receptor related role in spontaneous hypertension.

Animals↗
PubMed1988

Amino acid sequences of two nonspecific lipid-transfer proteins from germinated castor bean.

The amino acid sequence of two nonspecific lipid-transfer proteins (nsLTP) B and C from germinated castor bean seeds have been determined. Both the proteins consist of 92 residues, as for nsLTP previously reported, and their calculated Mr values are 9847 and 9593 for nsLTP-B and nsLTP-C, respectively. The sequences of nsLTP-B and nsLTP-C, compared to the known sequence of nsLTP-A from the same source, are 68% and 35% similar, respectively. No variation was found at the positions of the cysteine residues, indicating that they might be involved in disulfide bridges.

Amino Acid Sequence↗
PubMed1988

The structure of jack bean urease. The complete amino acid sequence, limited proteolysis and reactive cysteine residues.

The amino acid sequence of jack bean urease has been determined. The protein consists of a single kind of polypeptide chain containing 840 amino acid residues. The subunit relative molecular mass calculated from the sequence is 90,770, indicating that urease is composed of six subunits. Out of 25 histidine residues in urease, 13 were crowded in the region between residues 479 and 607, suggesting that this region may contain the nickel-binding site. Limited tryptic digestion cleaved urease at two sites, Lys-128 and Lys-662. Proteolytic products were not dissociated and retained full enzymatic activity. Five tryptic peptides containing the reactive cysteine residues were isolated and characterized with the aid of sulfhydryl-specific reagents, N-iodoacetyl-N'-(5-sulfo-1-naphthyl)ethylenediamine and N-(7-dimethylamino-4-methyl-3-coumarinyl)-maleimide. The reactive cysteine residues were located at positions 59, 207, 592, 663, and 824. The possibility that Cys-59, Cys-207, Cys-663, and Cys-824 are involved in the urease activity of the enzyme has been eliminated. Cys-592, which is essential for enzymatic activity, is located in the above-mentioned histidine-rich region.

Amino Acid Sequence↗
PubMed1988

On the activities of parathyroid hormone-like factor and transforming growth factors in extract of pancreatic cancer associated with humoral hypercalcemia of malignancy.

Extract of exocrine pancreatic cancer associated with humoral hypercalcemia of malignancy was examined for biological activities of PTH-like factor and transforming growth factor (TGFA and TGFB), both of which are possible causes of hypercalcemia. The crude extract had both PTH-like and TGF activities. On Bio Gel P-60 column chromatography, PTH-like and TGFA activities were eluted at around 10 kD, whereas TGFB activity was eluted at around void fractions, 10 kD and 6 kD. Liver extract, used as a control material, exhibited only TGFB activity at around 6 kD. CM-cellulose column chromatography of 10 kD fractions resulted in a subtle distinction between PTH-like activity and TGF activities. Further fractionation of the peak with PTH-like activity on reverse-phase high performance liquid chromatography separated PTH-like activity distinctly from TGFB activity. TGFA activity was lost through the procedure. It was concluded that the exocrine pancreatic cancer associated with hypercalcemia produced not only PTH-like activity but also TGFA and TGFB activities. Several chromatographic analyses suggested that PTH-like activity and at least TGFB activity stem from distinct molecules and that the PTH-like factor has no significant TGFB activity intrinsically.

Cell Line↗
PubMed1987

Location of the essential cysteine residue of jack bean urease.

Jack bean urease is inactivated by the modification of about one cysteine residue per subunit with N-ethylmaleimide (NEM) or other thiol titrant. The location of this cysteine residue was identified. After blocking the unessential thiol groups with NEM, the essential cysteine was labeled with N-(4-dimethylamino-3,5-dinitrophenyl)maleimide (DDPM). The DDPM-labeled protein was cleaved with cyanogen bromide and a DDPM-fragment was purified by gel filtration and ion exchange chromatography. Amino-terminal amino acid sequence of the DDPM-labeled fragment corresponded to that of a cyanogen bromide fragment of urease, VCHHLDREIPEDLAFAHSRIRKKTIAAEDVLNDIGAISIISSDSQAM. The second residue, Cys-592 in native urease, was labeled with DDPM. Fluoride ion, which competitively inhibits urease activity, protected urease from inactivation by NEM. The dissociation constant of fluoride ion obtained from the rate of inactivation with NEM was essentially identical to both the kinetically and spectrophotometrically determined dissociation constants. These suggest that the essential cysteine is at or near the active site.

Amino Acid Sequence↗