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Biomedical subjects

Y Takai

Publications and source records attributed to Y Takai.

At least 253 records · Page 14Linked to original sources
PubMed1995

Purification and characterization of REKS from Xenopus eggs. Identification of REKS as a Ras-dependent mitogen-activated protein kinase kinase kinase.

We have previously identified a protein factor, named REKS (Ras-dependent Extracellular signal-regulated kinase/Mitogen-activated protein kinase kinase (MEK) Stimulator), which is necessary for Ras-dependent MEK activation. In this study, we attempted to highly purify and characterize REKS. We have highly purified REKS by successive column chromatographies using a cell-free assay system in which REKS activates recombinant extracellular signal-regulated kinase 2 through recombinant MEK in a guanosine 5'-O-(thiotriphosphate) (GTP gamma S)-Ki-Ras-dependent manner. REKS formed a stable complex with GTP gamma S-Ras; REKS was coimmunoprecipitated with GTP gamma S-Ki-Ras or GTP gamma S-Ha-Ras, but not with GDP-Ki-Ras or GDP-Ha-Ras by an anti-Ras antibody. REKS was absorbed to a GTP gamma S-glutathione S-transferase (GST)-Ha-Ras-coupled glutathione-agarose column but not to a GDP-GST-Ha-Ras-coupled glutathione-agarose column and was coeluted with GTP gamma S-GST-Ha-Ras by reduced glutathione. The minimum molecular mass of REKS was estimated to be about 98 kDa on SDS-polyacrylamide gel electrophoresis. REKS phosphorylated this 98-kDa protein as well as recombinant MEK. REKS was not recognized by any of the anti-c-Raf-1, anti-Mos, and anti-mSte11 antibodies. These results indicate that REKS is a Ras-dependent MEK kinase.

Animals↗
PubMed1995

[Clinical evaluations of subtotal thyroidectomy for Graves' disease].

An attempt was made to determine the clinical evaluation of subtotal thyroidectomy in 58 patients with Graves' disease. The weight of the remnant thyroid gland was measured during the operations. Postoperative thyroid volume was measured by ultrasonography (thyroid volume = pi abc/6; a is length, b width, and c thickness). Postoperative thyroid function including serum TSH, free triiodothyronine (FT3), free thyroxine (FT4), thyroglobulin (Tg), TSH receptor antibodies (TRAb), and antimicrosomal antibodies (MCHA) was examined. Fifty-eight patients were divided into the three groups, according to postoperative thyroid function; 39 (67.2%) in remission, 7 (12.1%) with relapse and 12 (20.7%) in a hypothyroid state. Postoperative thyroid volume in patients with relapse was significantly (p < 0.05) greater than that in patients in remission and in a hypothyroid state. Remnant thyroid weight in patients with relapse was significantly (p < 0.05) heavier than that in patients in remission and in a hypothyroid state. However, there was no significant difference in an estimated total thyroid weight among the three groups. A significant correlation was noted between the remnant thyroid weight and the postoperative thyroid volume (R = 0.58, p < 0.001). On the other hand, there was a significant correlation between serum Tg level and the postoperative thyroid volume (R = 0.45, p < 0.01). Serum level of Tg in patients with relapse was significantly (p < 0.05) higher than that in patients in remission and in a hypothyroid state. The prevalence of negative MCHA in patients in remission appeared relatively higher than that in patients with relapse and in a hypothyroid state. In patients with a remnant thyroid weight of less than 4.0 g, there was no recurrence following subtotal thyroidectomy, but a higher prevalence of hypothyroid state (43%) was observed compared to those of larger remnant thyroid weight. In patients with a remnant thyroid weight of 4.0 to 6.0 g, there was a lower prevalence of recurrence (5.9%) compared to those with a larger remnant thyroid weight, while most patients showed remission (73.5%). On the other hand, the highest prevalence of recurrence (23.5%) was obtained in patients with a remnant thyroid weight of more than 6.0 g. These observations indicate that postopertive thyroid state does not depend upon an estimated total thyroid weight, but depends upon a remnant thyroid weight. In addition, a remnant thyroid weight is closely associated with thyroid volume measured by ultrasonography after subtotal thyroidectomy, which reflected the serum level of Tg.

Graves Disease↗
PubMed1995

Doc2: a novel brain protein having two repeated C2-like domains.

Two repeated C2-like domains interacting with Ca2+ and phospholipid are found in synaptotagmin and Rabphilin-3A which are implicated in neurotransmitter release. Here we have isolated a cDNA encoding a novel protein having two repeated C2-like domains from a human brain cDNA library. The isolated cDNA encodes a protein with 400 amino acids and a M(r) of 44,071. The purified recombinant protein indeed interacts with Ca2+ and phospholipid. We have named this protein Doc2 (Double C2). Doc2 is exclusively expressed in brain and is highly concentrated in the synaptic vesicle fraction. These results suggest that Doc2 is a novel brain protein and serves as a Ca2+ sensor in neurotransmitter release.

Adaptor Proteins, Signal Transducing↗
PubMed1995

Cytotoxic metabolites from a fungal adherent of a marine alga.

Leptosins G, G1, G2 and H, belonging to a series of dimeric epipolythiodioxopiperazines, were isolated from the mycelium of a strain of Leptosphaeria sp. stuck on the marine alga Sargassum tortile. Their stereostructures were elucidated by chemical and spectral evidence.

Antineoplastic Agents↗
PubMed1995

Rabphilin-3A binds to a M(r) 115,000 polypeptide in a phosphatidylserine- and Ca(2+)-dependent manner.

Rabphilin-3A is a putative target protein for Rab3A/Smg 25A, which is a member of the Ras-related small GTP-binding protein and implicated in neurotransmitter release from the synapse. Rabphilin-3A is composed of two functionally different domains: the N-terminal Rab3A-binding and the C-terminal phosphatidylserine- and Ca(2+)-binding domains. The C-terminal domain has two copies of an internal repeat that are homologous to the C2 domains of protein kinase C, synaptotagmin, and phospholipase A2 and C-gamma 1, which are known to bind phosphatidylserine and Ca2+. In this study, we attempted to identify the Rabphilin-3A-interacting molecule in bovine brain by use of an overlay assay technique. The 32P-labeled C-terminal fragment of Rabphilin-3A (281-704 amino acids) bound to a protein molecule with a M(r) of about 115 kDa which was immobilized on a nitrocellulose sheet. This protein was highly purified and characterized. The binding of the 32P-labeled C-terminal fragment to this protein was dependent on both phosphatidylserine and Ca2+, and inhibited by an excess amount of the C-terminal fragment and the C2 domain fragment (396-704 amino acids) but not by the N-terminal fragment (1-280 amino acids). These results indicate that Rabphilin-3A binds to a protein molecule with a M(r) of 115 kDa through the C2 domain in the presence of phosphatidylserine and Ca2+.

Adaptor Proteins, Signal Transducing↗
PubMed1995

Immunohistochemical evaluation of tenascin in benign eccrine gland tumors: comparison with laminin and fibronectin.

Surgically resected specimens of skin appendage tumors, spiradenoma, poroma of eccrine glands, syringoadenoma, syringoadenoma papilliferum and mixec tumor of skin were evaluated for tenascin, as well as the basement membrane-associated protein laminin, and the extracellular matrix protein fibronectin, using immunohistochemical methods. In normal sweat glands, tenascin was localized as a delicate, thin band in the immediate vicinity of the basement membrane and laminin was limited to the basement membrane. Tenascin in syringoadenoma was also expressed linearly adjacent to the basement membrane. Mixed tumors of skin showed a characteristic distribution of tenascin in the modified myoepithelial cell area adjacent to the tumor epithelia with a heterogeneous pattern, and particularly strong expression in the hyalinous and chondroid areas. In addition, the localization of laminin in the basement membranes of tumors, intracellular immunoreactivity was observed in syringoadenoma, syringocystadenoma papilliferum, and mixed tumor of skin. Fibronectin reactivity was visualized as a diffuse staining in the tumor stroma of eccrine poroma but its expression was restricted in eccrine spiradenoma. The present study suggests a heterogeneity in the distribution of basement membrane and extracellular matrix proteins in skin appendage tumors and elucidation of their functional roles would assist in understanding the changes in the supporting tumor stroma following neoplastic changes.

Cell Adhesion Molecules, Neuronal↗
PubMed1995

Expression of tenascin in odontogenic tumours.

We investigated the expression of tenascin in a series of odontogenic tumours (n = 63) of epithelial and epithelial-ectomesenchymal origin by using immunohistochemical methods. A heterogeneity of expression of tenascin was observed in odontogenic tumours. The heterogeneity was most prominent in odontogenic tumours not forming calcified tissues. In these ameloblastomas and adenomatoid odontogenic tumours, tenascin was mainly localised at the epithelial tumour cell-mesenchymal tissue interface. In the calcifying epithelial odontogenic tumour, ameloblastic fibroma and odontoma, a widespread stromal immunoreactivity was observed which was, however, unreactive in the calcified masses. The stellate reticulum-like cells and granular cells of ameloblastoma also showed a positive immunoreactivity for tenascin. The results of the present study suggest that expression of tenascin in the stromal tissue of odontogenic tumours differs according to the potential of forming calcified masses by the tumour cells irrespective of tumour cell morphology.

Ameloblastoma↗
PubMed1995

Rho as a regulator of the cytoskeleton.

Many cell functions, including maintenance of morphology, aggregation, motility, membrane ruffling, smooth-muscle contraction, cytokinesis in mammals and bud formation in yeast, are regulated through the dynamic reorganization of actin filaments. Although it has long been known that Ca2+ is a key regulator of the cytoskeleton, evidence is now accumulating that Rho, a Ras-related small GTP-binding protein, is another important regulator of these actin-dependent cell functions.

Acute-Phase Proteins↗
PubMed1995

Diagnostic criteria for neoplastic myoepithelial cells in pleomorphic adenomas and myoepitheliomas. Immunocytochemical detection of muscle-specific actin, cytokeratin 14, vimentin, and glial fibrillary acidic protein.

OBJECTIVE: Markers for normal salivary gland myoepithelium were used to determine the extent of their expression in the neoplastic myoepithelial (nonluminal) cells of pleomorphic adenomas and then in the tumor cells in myoepitheliomas and to gather information necessary to establish diagnostic criteria, especially muscle actin expression, for myoepitheliomas. STUDY DESIGN: Methanol/acetic acid-fixed and paraffin-embedded tissue was used to immunohistochemically study expression of intermediate and smooth-muscle actin filaments in nonluminal cells in 14 pleomorphic adenomas and to compare this to their expression in five myoepitheliomas. RESULTS: In routine histologic sections, the morphologic variants of nonluminal tumor cells--spindle, stellate, polygonal, angular, and plasmacytoid--in pleomorphic adenoma mirror the spectrum of tumor cells in myoepitheliomas. Immunocytochemical similarities are also apparent. Two specific markers for myoepithelial cells in the normal salivary gland, muscle-specific actin and cytokeratin 14, were both variably, independently, and never uniformly expressed in nonluminal cells of pleomorphic adenoma and tumor cells in myoepitheliomas regardless of their morphology. Cytokeratin 14 in addition labels basal cells of excretory ducts. Both muscle-specific actin and cytokeratin 14 preferentially localized to single layers of periductal cells in pleomorphic adenomas, angular, polygonal, and plasmacytoid cells preferentially expressed cytokeratin 14. Similar patterns were noted in the three myoepitheliomas with reasonable expression of the two markers. Only isolated single cells or small groups of plasmacytoid cells in four pleomorphic adenomas with a significant component of these cells and the two plasmacytoid myoepitheliomas immunostained for muscle-specific actin and cytokeratin 14. In both tumor types, vimentin was nearly uniformly expressed in nonluminal tumor cells of all morphologic types, including plasmacytoid cells. CONCLUSIONS: The range and transition of morphology of nonluminal cells in pleomorphic adenomas is reflected in myoepitheliomas. Incomplete or absent expression of the myoepithelial/basal cell markers, muscle-specific actin, and cytokeratin 14, and the general expression of vimentin is common to both tumors. Because these findings apply to the majority of plasmacytoid cells in pleomorphic adenomas, tumor cells with a similar morphology and immunoprofile are to be expected in myoepitheliomas; the term plasmacytoid myoepitheliomas is thus appropriate regardless of the presence or absence of muscle-specific actin.

Actins↗
PubMed1995

Differential targeting of protein kinase C and CaM kinase II signalings to vimentin.

Hydrolysis of inositol phospholipids by receptor stimulation activates two separate signaling pathways, one leading to the activation of protein kinase C (C kinase) via formation of diacylglycerol. The other is the inositol trisphosphate (IP3)/Ca2+ pathway and a major downstream kinase which is activated is Ca2+/calmodulin-dependent protein kinase II (CaM kinase II). To examine signaling pathways of C kinase and CaM kinase II to the cytoskeletal protein vimentin, we prepared monoclonal antibodies YT33 and MO82 which recognize the phosphorylation state of vimentin by C kinase and by CaM kinase II, respectively. Ectopic expression of constitutively active C kinase or CaM kinase II in primary cultured astrocytes by microinjection of the corresponding expression vectors induced phosphorylation of vimentin at each specific phosphorylation site, followed by reorganization of vimentin filament networks. In contrast, simultaneous activation of C kinase and CaM kinase II by inositol phospholipid hydrolysis with receptor stimulation led to an exclusive phosphorylation of vimentin at the CaM kinase II site, not at the site of C kinase. These results indicate that the intracellular targeting of C kinase and CaM kinase II signalings to vimentin is regulated separately, under physiological conditions.

Amino Acid Sequence↗