PubMed Health⌕ Search

Biomedical subjects

D Watanabe

Publications and source records attributed to D Watanabe.

At least 37 records · Page 2Linked to original sources
PubMed2000

Identification of nuclear export signal in UL37 protein of herpes simplex virus type 2.

The UL37 gene of herpes simplex virus (HSV) encodes a 120-kDa phosphoprotein associated with the virion. In this study, we have generated a rabbit polyclonal antiserum against HSV-2 UL37 protein, and examined its intracellular localization by immunofluorescence study. In infected cells, specific fluorescence was detectable in the perinuclear region. In transfected cells, UL37 protein was observed mainly in the cytoplasm. Transfection assays of deletion mutants of UL37 protein suggested that the leucine rich region (LRR) containing amino acids 263-273 may be important for cytoplasmic localization. Deletion of the LRR or substitution of the leucine residues resulted in nuclear remaining of UL37 protein. Moreover, the LRR could export green fluorescent protein (GFP) to the cytoplasm as a fusion protein and this export was blocked by leptomycin B treatment, indicating that the LRR acted as a nuclear export signal. These results suggest that UL37 protein fulfills a role as a shuttle between the nucleus and the cytoplasm through the LRR.

Active Transport, Cell Nucleus↗
PubMed2000

Synaptic integration mediated by striatal cholinergic interneurons in basal ganglia function.

The physiological role of striatal cholinergic interneurons was investigated with immunotoxin-mediated cell targeting (IMCT). Unilateral cholinergic cell ablation caused an acute abnormal turning behavior. These mice showed gradual recovery but displayed abnormal turning by both excess stimulation and inhibition of dopamine actions. In the acute phase, basal ganglia function was shifted to a hyperactive state by stimulation and suppression of striatonigral and striatopallidal neurons, respectively. D1 and D2 dopamine receptors were then down-regulated, relieving dopamine-predominant synaptic perturbation but leaving a defect in controlling dopamine responses. The acetylcholine-dopamine interaction is concertedly and adaptively regulated for basal ganglia synaptic integration.

Acetylcholine↗
PubMed2000

A juvenile hormone-repressible transferrin-like protein from the bean bug, Riptortus clavatus: cDNA sequence analysis and protein identification during diapause and vitellogenesis.

We found several juvenile hormone-responsive cDNAs in the bean bug, Riptortus clavatus, by using mRNA differential display (Hirai et al., 1998). One of them, a juvenile hormone-repressible cDNA, JR-3, was cloned, sequenced, characterized and identified as a transferrin (RcTf). RcTf cDNA encoded 652 amino acids with a calculated molecular weight of 71,453 Da. The deduced amino acid sequence showed significant homology with the transferin genes of several insects, Manduca sexta (43% identity), Blaberus discoidalis (43%), Aedes aegypti (43%), Drosophila melanogaster (36%), Sarcophaga peregrina (36%) and the human (25%). Antiserum was prepared by using recombinant RcTf protein expressed in Escherichia coli as an antigen. The antiserum reacted specifically with both the recombinant protein and the native protein from the bugs, with sizes of 70 and 75 kDa, respectively. The 75 kDa protein was partially purified from hemolymph of diapausing female bugs and the first ten amino acids were found to be identical to that of RcTf cDNA, indicating that the 75 kDa protein is RcTf. The tissue distribution of RcTf in the bug was examined by Western blot analysis. In diapausing animals, RcTf was detected in the fat body, hemolymph and ovary but not in the gut. In the post-diapause stage, RcTf was also detected in eggs, in addition to the fat body and ovary. These results indicate that RcTf is incorporated into the oocytes during vitellogenesis, and suggest that it may provide iron for the developing embryos.

Amino Acid Sequence↗
PubMed2000

Corneal infection of herpes simplex virus type 2--induced neuronal apoptosis in the brain stem of mice with expression of tumor suppressor gene (p53) and transcription factors.

To understand the mechanism of neuronal apoptosis induced by herpes simplex virus (HSV) infection in vivo, the distribution of viral antigen, the appearance of apoptotic bodies, and the expressions of the tumor suppressor gene p53 and several transcription factors such as c-fos, c-jun and NF-kappaB were examined immunohistochemically and histopathologically after corneal infection of mice with HSV type 2 strain 186. Five days after HSV infection, viral antigen was diffusely detected in the corneal epithelium, the trigeminal ganglion and the pars caudalis of the spinal trigeminal nucleus. Neuronal apoptosis was observed in the brain stem ipsilateral to the HSV-infected side with the immunoreactivities of c-fos, c-jun, NF-kappaB and p53. Dual-labeling immunohistochemical studies revealed that almost all of the viral antigen-positive neurons and glia in the brain stem also showed p53 immunoreactivity. On the other hand, no neuronal apoptosis but only with the expression of c-jun was found in the trigeminal ganglion. Our results suggest that the different expression of transcription factors between the brain stem and the trigeminal ganglion may influence the neuronal apoptosis induced by HSV infection.

Afferent Pathways↗
PubMed2000

Herpes simplex virus UL17 protein is associated with B capsids and colocalizes with ICP35 and VP5 in infected cells.

A previous study using a mutant lacking the UL17 gene has suggested that the UL17 protein of herpes simplex virus type 1 (HSV-1) is required for the cleavage/packaging of viral DNA. In this study, we have raised a rabbit polyclonal antiserum which specifically reacted with the UL17 protein which has an apparent molecular mass of 78-kDa in the lysates of HSV types 1- and 2-infected Vero cells. Western blot analysis of intracellular capsids demonstrates that the UL17 protein was associated with B and C capsids. Indirect immunofluorescence studies reveal that it colocalized with the major capsid protein VP5 and the scaffolding protein ICP35 within the nucleus. These results suggest that the association of the UL17 protein with immature B-type capsids is important for its role in cleavage/packaging.

Animals↗
PubMed2000

Descending necrotizing mediastinitis due to odontogenic infections.

OBJECTIVE: Acute purulent mediastinitis caused by oropharyngeal infection is termed descending necrotizing mediastinitis. Such infections usually have a fulminate course, leading to sepsis and frequently to death. The purpose of this study is to show the importance of early diagnosis, aggressive surgical intervention, and optimal antibiotics chemotherapy in controlling this fatal infectious disease. STUDY DESIGN: Two patients with descending necrotizing mediastinitis due to odontogenic infection who were treated at our institution are described. RESULTS: Both patients survived. CONCLUSIONS: From the patients, 23 different aerobic and anaerobic bacteria were isolated. All of the isolates were susceptible to carbapenem. Early evaluation by means of cervicothoracic computed tomography scanning was extremely useful for diagnosis and surgical planning. Knowledge of anatomic pathways from the mouth to the mediastinum is essential. We believe that tracheostomy is not always necessary. In both of the cases presented, mediastinal drainage was completed through use of a transcervical approach. However, a more aggressive drainage including tracheostomy might be necessary when the infection extends below the carina.

Acute Disease↗
PubMed2000

The imprinted antisense RNA at the Igf2r locus overlaps but does not imprint Mas1.

The gene encoding the insulin-like growth-factor type-2 receptor (Igf2r) is maternally expressed and imprinted. A CpG island in Igf2r intron 2 that carries a maternal-specific methylation imprint was shown in a transgenic model to be essential for Igf2r imprinting and for the production of an antisense RNA from the paternal allele. We report here that the endogenous region2 is the promoter for this antisense RNA (named Air, for antisense Igf2r RNA) and that the 3' end lies 107,796 bp distant in an intron of the flanking, but non-imprinted, gene Mas1.

Base Sequence↗
PubMed2000

Malignant melanoma in the thymus.

A case of malignant melanoma in the thymus is reported. Diagnostic imaging demonstrated a left anterior mediastinal mass in a patient with giant pigmented nevus without malignant change. Histologic and cytologic specimens obtained from the tumor revealed that the tumor was malignant melanoma. Surgery revealed malignant melanoma in the left lobe of the thymus. Many cell nests of pigmented nevi were observed throughout the thymus. The malignant melanoma was thought to have originated from the nevocellular nevus in the thymus. This is the first report of malignant melanoma in the thymus.

Adult↗
PubMed1999

Structure and function of the human transcription elongation factor DSIF.

5,6-Dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) is a classic inhibitor of transcription elongation by RNA polymerase II (pol II). We have previously identified and purified a novel transcription elongation factor, termed DSIF (for DRB sensitivity-inducing factor), that makes transcription sensitive to DRB. DSIF is composed of 160- and 14-kDa subunits, which are homologs of the Saccharomyces cerevisiae transcription factors Spt5 and Spt4. DSIF may either repress or stimulate transcription in vitro, depending on conditions, but its physiological function remains elusive. Here we characterize the structure and function of DSIF p160. p160 is shown to be a ubiquitous nuclear protein that forms a stable complex with p14 and interacts directly with the pol II largest subunit. Mutation analysis of p160 is used to identify structural features essential for its in vitro activity and to map the domains required for its interaction with p14 and pol II. Finally, a p160 mutant that represses DSIF activity in a dominant-negative manner is identified and used to demonstrate that DSIF represses transcription from various promoters in vivo.

Amino Acid Sequence↗
PubMed1999

The role of polymorphonuclear leukocyte infiltration in herpes simplex virus infection of murine skin.

We undertook the present study to investigate the role of polymorphonuclear leukocytes (PMN) in defending skin against herpes simplex virus (HSV) infection. For this purpose, we established a mouse model of cutaneous HSV infection. The hind limb footpad skin of 4-week-old ICR mice was abraded linearly once with a feather edge file and infected with various strains of HSV with different virulence. In uninfected control mice, PMN appeared at the abraded skin lesion within 24 h, and were eliminated from the epidermis after 3 days. Mice inducted with a highly virulent strain of HSV demonstrated wide and severe erythematous lesions of the footpad skin and histologically, virus antigen-positive ballooning degenerated keratinocytes were observed. However, in infections with attenuated strains of HSV, the epidermis was regenerated and a viral antigen was discharged within 5 days, together with any infiltrated PMN. Macrophages and NK cells numbered less than PMN. In mice treated with anti-PMN antiserum before HSV infection, PMN infiltration was significantly suppressed 1 day after infection, and these animals developed a severe cutaneous disease even if infected with an attenuated virus. These results indicate the importance of PMN in the control of HSV cutaneous infections, especially in the primary infectious phase.

Animals↗
PubMed1999

US3 protein kinase of herpes simplex virus type 2 plays a role in protecting corneal epithelial cells from apoptosis in infected mice.

To clarify the biological role of US3 protein kinase of herpes simplex virus type 2 (HSV-2) in vivo, the expression of the viral antigen, the appearance of apoptotic bodies and DNA fragmentation were examined immunohistologically after corneal infection of mice with three different kinds of HSV-2 strain 186: the wild-type virus, a US3-deficient mutant (L1BR1) and its revertant (L1B-11). In both wild-type 186- and L1B-11-infected mice, viral antigen was diffusely found in the corneal epithelium; no apoptotic changes were detected in the epithelial cells. Whereas, in L1BR1-infected mice, HSV-immunoreactivity was localized around the virus-inoculated sites, and a large number of apoptotic bodies were observed in the corneal epithelium with dual-positive reactions for both HSV-immunostaining and TUNEL staining. These results suggest that the US3 protein kinase plays an important role in protecting HSV-2-infected cells from apoptotic death in vivo.

Animals↗
PubMed1999

Keratoacanthoma centrifugum marginatum arising from a scar after skin injury.

Among the variants of solitary keratoacanthoma, keratoacanthoma centrifugum marginatum (KCM) is characterized by the lack of a tendency toward spontaneous remission and by continuous centrifugal spread. We describe a case of KCM arising from the scar after an old skin injury. The lesion appeared on the dorsum of the right hand, grew peripherally for 30 months, and became a tumor with a multinodular margin and central atrophy. A biopsy specimen from the edge of the tumor showed features resembling typical solitary keratoacanthoma.

Aged↗
PubMed1998

Evidence that P-TEFb alleviates the negative effect of DSIF on RNA polymerase II-dependent transcription in vitro.

Recently, a positive and a negative elongation factor, implicated in 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) inhibition of transcription elongation, has been identified. P-TEFb is a positive transcription elongation factor and the DRB-sensitive kinase that phosphorylates the C-terminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II). PITALRE, a member of the Cdc2 family of protein kinases, is the catalytic subunit of P-TEFb. DSIF is a human homolog of the yeast Spt4-Spt5 complex and renders elongation of transcription sensitive to DRB. DRB sensitivity-inducing factor (DSIF) binds to RNA Pol II and may directly regulate elongation. Here we show a functional interaction between P-TEFb and DSIF. The reduction of P-TEFb activity induced by either DRB, antibody against PITALRE, or immunodepletion resulted in a negative effect of DSIF on transcription. DSIF acts at an early phase of elongation, and the prior action of P-TEFb makes transcription resistant to DSIF. The state of phosphorylation of CTD determines the DSIF-RNA Pol II interaction, and may provide a direct link between P-TEFb and DSIF. Taken together, this study reveals a molecular basis for DRB action and suggests that P-TEFb stimulates elongation by alleviating the negative action of DSIF.

Cyclin-Dependent Kinase 9↗
PubMed1998

Ablation of cerebellar Golgi cells disrupts synaptic integration involving GABA inhibition and NMDA receptor activation in motor coordination.

The role of inhibitory Golgi cells in cerebellar function was investigated by selectively ablating Golgi cells expressing human interleukin-2 receptor alpha subunit in transgenic mice, using the immunotoxin-mediated cell targeting technique. Golgi cell disruption caused severe acute motor disorders. These mice showed gradual recovery but retained a continuing inability to perform compound movements. Optical and electrical recordings combined with immunocytological analysis indicated that elimination of Golgi cells not only reduces GABA-mediated inhibition but also attenuates functional NMDA receptors in granule cells. These results demonstrate that synaptic integration involving both GABA inhibition and NMDA receptor activation is essential for compound motor coordination. Furthermore, this integration can adapt after Golgi cell elimination so as not to evoke overexcitation by the reduction of NMDA receptors.

Animals↗
PubMed1998

Glutamate receptors: brain function and signal transduction.

Glutamate receptors are important in neural plasticity, neural development and neurodegeneration. N-methyl-d-aspartate (NMDA) receptors and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/kainate receptors act as glutamate-gated cation channels, whereas metabotropic receptors (mGluRs) modulate the production of second messengers via G proteins. Molecular studies from our and other laboratories indicated that NMDA receptors and mGluRs exist as multiple subunits (NMDAR1 and NMDAR2A-2D) and multiple subtypes (mGluR1-mGluR8). In light of the molecular diversity of glutamate receptors, we explored the function and intracellular signaling mechanisms of different members of glutamate receptors. In the visual system, retinal bipolar cells receive glutamate transmission from photoreceptors and contribute to segregating visual signals into ON and OFF pathways. The molecularly cloned mGluR6 is restrictedly expressed at the postsynaptic site of ON-bipolar cells in both rod and cone systems. Gene targeting of mGluR6 results in a loss of ON responses without changing OFF responses and severely impairs detecting visual contrasts. Since AMPA receptors mediate OFF responses in OFF-bipolar cells, two distinct types of glutamate receptors effectively operate for ON and OFF responses. mGluR1 and mGluR5 are both coupled to inositol triphosphate (IP3)/calcium signal transduction with an identical agonist selectivity. Single-cell intracellular calcium ([Ca2+]i) recordings indicated that glutamate evokes a non-oscillatory and oscillatory [Ca2+]i response in mGluR1-expressing and mGluR5-expressing cells, respectively. This difference results from a single amino acid substitution, aspartate of mGluR1 or threonine of mGluR5, at the G protein-interacting carboxy-terminal domains. Protein kinase C phosphorylation of the threonine of mGluR5 is responsible for inducing [Ca2+]i oscillations in mGluR5-expressing cells and cultured glial cells. Thus, the two closely related mGluR subtypes mediate diverging intracellular signaling in glutamate transmission.

Animals↗
PubMed1998

Nucleotide sequence of vitellogenin mRNA in the bean bug, Riptortus clavatus: analysis of processing in the fat body and ovary.

The bean bug, Riptortus clavatus, has immunologically distinct yolk proteins, vitellin (Vn)-1 and-2 and their precursors, vitellogenin (Vg)-1 and-2. We have cloned the full nucleotide sequence of Vg-1 cDNA. The deduced amino acid sequence has some similarities to other insect Vgs. It contains two polyserine regions, which are characteristic of other Vgs. Vg-1 mRNA appeared after treatment with the juvenile hormone analogue, methoprene, implying transcriptional regulation. We found four enzymatic cleavage sites in the Vg molecule. Two of them match the consensus for dibasic processing endoprotease, which is also conserved in processing sites for other insect Vgs. We showed that the processing at each site was incomplete, and this resulted in production of more than the five polypeptides which would be expected from four processing sites in the molecule. The physiological significance of multiple polypeptides in insect Vgs is still unclear.

Amino Acid Sequence↗
PubMed1998

Three new manzamine congeners from amphimedon sponge

Three new manzamine congeners (1-3) have been isolated from the Okinawan marine sponge Amphimedon sp., and the structures, including relative stereochemistries, were elucidated on the basis of spectroscopic data. Absolute configuration at C-16 of 1 was determined by modified Mosher's method.

Journal Article↗