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S Shiosaka

Publications and source records attributed to S Shiosaka.

At least 37 records · Page 2Linked to original sources
PubMed1999

cDNA cloning and expression of a novel serine protease in the mouse brain.

A cDNA for a novel serine protease, termed brain type granzyme K (B-GRK) was cloned from the mouse brain. The cDNA codes a protein similar to granzyme K (GRK) but completely different at the N-terminus. Genomic Southern and PCR analysis of the gene suggests B-GRK is the alternative transcription form of GRK. B-GRK and GRK have a different organ-specific expression pattern: B-GRK is expressed in the brain, while GRK is expressed in the spleen. The recombinant fusion protein was detected in the neuro2a cells transfected with a plasmid containing B-GRK sequence. The mRNA for B-GRK/GRK was detected in cerebral cortex, hippocampus and diencephalon of the mouse brain. In situ hybridization for B-GRK/GRK revealed that several regions in the forebrain and hypothalamus express the mRNA. Developmental analysis showed that in the prenatal stage, the mRNA was expressed also in pituitary and pineal body in addition to the brain.

Amino Acid Sequence↗
PubMed1999

Injury induces neuropsin mRNA in the central nervous system.

We have shown that neuropsin is expressed in the neurons of the limbic system in the adult mouse. After the central nervous system was injured by incision or intraperitoneal kainate injection, neuropsin mRNA was induced in the peri-lesioned region. The cells in which neuropsin mRNA was induced were localized mainly in axon fiber pathways and closely associated to proteolipid protein (PLP) mRNA expressing oligodendrocytes.

Animals↗
PubMed1999

Crystal structure of neuropsin, a hippocampal protease involved in kindling epileptogenesis.

Neuropsin is a novel serine protease, the expression of which is highly localized in the limbic areas of the mouse brain and which is suggested to be involved in kindling epileptogenesis and hippocampal plasticity. The 2.1-A resolution crystal structure of neuropsin provides the first three-dimensional view of one of the serine proteases highly expressed in the nervous system, and reveals a serine protease fold that exhibits chimeric features between trypsin and nerve growth factor-gamma (NGFgamma), a member of the kallikrein family. Neuropsin possesses an N-glycosylated "kallikrein loop" but forms six disulfide bonds corresponding to those of trypsin. The ordered kallikrein loop projects proline toward the active site to restrict smaller residues or proline at the P2 position of substrates. Loop F, which participates in forming the S3/S4 sites, is similar to trypsin rather than NGFgamma. The unique conformations of loops G and H form an S1 pocket specific for both arginine and lysine. These characteristic loop structures forming the substrate-binding site suggest the novel substrate specificity of neuropsin and give a clue to the design of its specific inhibitors.

Amino Acid Sequence↗
PubMed1999

Effect of 12-O-tetradecanoyl-phorbol ester and incisional wounding on neuropsin mRNA and its protein expression in murine skin.

The expression of neuropsin mRNA in vivo in mouse skin was examined by in situ hybridization and Northern blotting under stimulated conditions. Two kinds of epidermal stimuli, a topical application of a chemical tumor promoter and incisional wounding, were used. A single topical application of 12-O-tetradecanoyl-phorbol 13-acetate induced epidermal hyperplasia and simultaneously induced an extensive increase in neuropsin mRNA in the suprabasal cells. A full-thickness skin incision also induced a profound increase in neuropsin mRNA in the suprabasal cells surrounding the wound but not in actively proliferating basal cells. The increases in neuropsin mRNA occurred rather late and were limited to the site of drug application or around the incision. Interestingly, neuropsin mRNA was not expressed in the epithelial tongue migrating toward the wound during re-epithelialization. Thus, neuropsin might participate in accelerated epidermal differentiation rather than in the proliferation or migration of keratinocytes in the wound.

Animals↗
PubMed1999

High expression of alternative transcript of granzyme M in the mouse retina.

We have isolated cDNAs to two transcripts, granzyme M and alternative granzyme M mRNA from the mouse eye. Analysis of genomic DNA revealed these transcripts were derived from alternative transcription initiations. Northern blot analysis and reverse transcription-polymerase chain reactions revealed that both transcripts were expressed in the eye, though the alternative form was the major type. In situ hybridization studies demonstrated that alternative granzyme M mRNA localized exclusively in the photoreceptor cells in the retina and expressed only after the opening of the eye, suggesting that these transcripts are related to the maintenance of the retinal structure or functions of matured photoreceptor cells rather than the development or differentiation of retinal cells.

Alternative Splicing↗
PubMed1999

N-glycan structures of murine hippocampus serine protease, neuropsin, produced in Trichoplusia ni cells.

N-glycans of neuropsin (serine protease in the murine hippocampus) expressed in Trichoplusia ni cells were released from the glycopeptides by digestion with glycoamidase A (from sweet almond), and the reducing ends of the oligosaccharides were reductively aminated with 2-aminopyridine. The derivatized N-glycans were separated and structurally identified by a two dimensional high-performance liquid chromatography (HPLC) mapping technique on two kinds of HPLC columns. Fourteen different major N-glycan structures were identified, of which 6 were high-mannose type (9.1%), and the remaining 8 were paucimannosidic type. The presence of insect specific N-glycan structures containing both alpha1,3- and alpha1,6- di-fucosylated innermost N-acetylglucosamine residue (23.3%), as below, was also confirmed by 600 MHz 1H-NMR spectroscopy.

Animals↗
PubMed1999

Plasticity-related serine proteases in the brain (review).

Serine proteases exert a variety of functions in the body; food digestion, regulation of other proteins and modification of extracellular matrix. Cumulative evidence has shown the importance of serine proteases in the nervous system as well. It has been shown that three serine proteases, thrombin, plasminogen activators and neuropsin, have functional roles in neural plasticity. Most of the actions of thrombin are thought to be mediated by its specific receptors. Thrombin reverses neurite outgrowth of serum-deprived neuroblastoma cells, and induces protective and apoptotic effects on neurons and glial cells depending on concentration and time. Tissue-type and urokinase-type plasminogen activators (tPA and uPA) distribute broadly in the brain. tPA and uPA exert a variety of functions during development. These proteases also function in long-term potentiation and kindling formation. Furthermore, tPA is essential to excitotoxic neuronal cell death. Neuropsin is a serine protease expressed in the limbic system of the brain. Kindling induced neuropsin mRNA and protein expression and anti-neuropsin antibody ameliorates kindling epilepsy. The possible roles of these proteases in neural plasticity are reviewed here.

Animals↗
PubMed1998

Molecular cloning and distinct developmental expression pattern of spliced forms of a novel zinc finger gene wiz in the mouse cerebellum.

In the course of a study conducted to identify the mouse homologue of Drosophila eyes absent (eya), we isolated a novel mouse cDNA fragment which show little homology to eya but encodes a protein with Krüppel (C2H2)-type zinc finger motifs. By further screening using this cDNA fragment as a probe, we obtained the short and long forms of full-length cDNAs, which were apparently alternatively spliced products from one gene. Since both mRNAs encode proteins with widely-interspaced zinc finger motifs, we termed this gene wiz and refer to the short and long wiz transcripts as wizS and wizL, respectively. In situ hybridization studies using the probe against the region common to wizS and wizL showed that these mRNAs were expressed abundantly in the granule cell layers of the mouse cerebellum, the olfactory bulb, and the dentate gyrus, whereas the same technique using the probe against only wizL could not detect positive signals in the developing cerebellum, indicating that there is no expression of wizL mRNA there. Northern blot and in situ hybridization analyses demonstrated that the extracerebellar regions expressed both wizS and wizL mRNAs from the midgestational period to adulthood. The finding that two types of wiz transcripts (wizS and wizL) are expressed with different developmental patterns might indicate separate transcription functions in the cerebellar granule cells and the extracerebellar regions.

Alternative Splicing↗
PubMed1998

cDNA cloning and expression of a novel serine protease, TLSP.

A cDNA for a putative novel serine protease, TLSP, was cloned from human hippocampus cDNA with polymerase chain reaction based strategies. The putative amino acid sequence of TLSP is similar to the trypsin-type serine proteases. TLSP mRNA is expressed in keratinocytes. Overexpressed TLSP protein in neuro2a cells was detected in culture medium.

Amino Acid Sequence↗
PubMed1998

Sequence analysis and expression of human neuropsin cDNA and gene.

Neuropsin is a serine protease which is thought to function in a variety of tissues including the brain and skin. This protease has been shown to have important roles in neural plasticity in mice. Here we have cloned a cDNA and analyzed the gene for human neuropsin by polymerase chain reaction-based strategies. The cDNA had 72% identity to mouse neuropsin. The deduced amino acid sequence showed 72% identity to mouse neuropsin. Key amino acid residues for the enzyme activity and all cysteine residues were conserved between human and mouse neuropsin. The gene for human neuropsin had six exons and five introns, and the gene organization is similar to trypsin-type serine proteases. The mRNA was expressed in primary cultures of keratinocytes.

Amino Acid Sequence↗
PubMed1998

Characterization of recombinant and brain neuropsin, a plasticity-related serine protease.

Activity-dependent changes in neuropsin gene expression in the hippocampus implies an involvement of neuropsin in neural plasticity. Since the deduced amino acid sequence of the gene contained the complete triplet (His-Asp-Ser) of the serine protease domain, the protein was postulated to have proteolytic activity. Recombinant full-length neuropsin produced in the baculovirus/insect cell system was enzymatically inactive but was readily converted to active enzyme by endoprotease processing. The activational processing of prototype neuropsin involved the specific cleavage of the Lys32-Ile33 bond near its N terminus. Native neuropsin that was purified with a purity of 1,100-fold from mouse brain had enzymatic characteristics identical to those of active-type recombinant neuropsin. Both brain and recombinant neuropsin had amidolytic activities cleaving Arg-X and Lys-X bonds in the synthetic chromogenic substrates, and the highest specific activity was found against Boc-Val-Pro-Arg-4-methylcoumaryl-7-amide. The active-type recombinant neuropsin effectively cleaved fibronectin, an extracellular matrix protein. Taken together, these results indicate that this protease, which is enzymatically novel, has significant limbic effects by changing the extracellular matrix environment.

Amino Acid Sequence↗
PubMed1998

Differentially expressed olfactomedin-related glycoproteins (Pancortins) in the brain.

Messenger RNA differential display is conducted to search for genes that are expressed in a region-specific pattern in the rodent brain. Eleven novel gene fragments are isolated. One of these genes which we call pancortin, based on its predominant mRNA expression in the cerebral cortex of the adult, is studied. These pancortin cDNA clones are grouped into four different types of cDNA, designated as pancortin-1 to -4. All pancortin cDNAs share a common sequence in the middle of their structure, having two alternative sequences at both 5'- and 3'-ends, respectively. Deduced amino acid sequence shows that all pancortins have sequences of hydrophobic amino acids at N-terminus and no obvious membrane spanning regions. In situ hybridization histochemistry using oligonucleotide probes specific for 5'- and 3'-end variable parts has revealed that these four pancortin mRNAs are expressed differentially in the adult rodent brain. Robust expression of pancortin-1 and -2 mRNA is observed in the cerebral cortex (including the hippocampus and the olfactory bulb). However, little of pancortin-3 and -4 mRNA is observed there. In the cortex, some neurons are stained by an antibody raised against Pancortin. Immuno-electron microscopic study has revealed that Pancortin-like immunoreactive products are localized mainly in the endoplasmic reticulum and not in the Golgi apparatus indicating that Pancortins are the endoplasmic reticulum-anchored proteins. Our results suggest that each Pancortin is differentially regulated and may perform different functions in the brain.

Amino Acid Sequence↗
PubMed1998

Blockade of neuropsin, a serine protease, ameliorates kindling epilepsy.

The behavioural and electrographical abnormalities associated with seizures in epileptic (kindled) mice correspond with those of human epilepsy. In kindled mice, neuropsin was markedly increased in the hippocampus and cerebral cortices. A single intraventricular injection of monoclonal antibodies specific to neuropsin reduced or eliminated the epileptic pattern noted on electroencephalograms and, as a result markedly inhibited the progression of kindling. Therefore, neuropsin appears to be a key protein controlling pathogenic events in the hippocampus, and thus neuropsin inhibitors might be useful for treatment of epilepsy.

Animals↗
PubMed1998

Expression of neuropsin in the keratinizing epithelial tissue-immunohistochemical analysis of wild-type and nude mice.

Neuropsin is a trypsin-type serine protease that was first cloned from the mouse brain as a factor related to neural plasticity. Subsequent in situ hybridization histochemical analysis indicated a broad localization of its mRNA throughout the whole body, although the details remain obscure. In this study, we showed that neuropsin immunoreactivity is localized in the keratinized stratified epithelia of the mouse epidermis, hair, tongue, palate, nasal cavity, pharynges, esophagus, and forestomach. In the skin and mucous membranes, neuropsin immunoreactivity was found in the stratum spinosum and the stratum granulosum. The immunoreactivity in the former sublayer was mainly present in the cytoplasm, but that in the latter sublayer was exclusively present in the intercellular space or on the outer surface of the cell membrane and thus exhibited a lamellar-like peripheral distribution. During development, the appearance of neuropsin immunoreactivity in the various epithelia was found at embryonic days 14.5-15.5, prior to formation of the stratum corneum. More extensive expression of neuropsin immunoreactivity was found in the nude mouse skin and mucous membranes than in wild-type mice. Because the nude mouse is characterized by genetic impairment of keratinization, such abnormal neuropsin expression might be caused or affected by this impairment. Therefore, neuropsin, an extracellular serine protease, is suggested to be involved in keratinization in the stratified epithelia.

Animals↗
PubMed1998

Expression of neuropsin mRNA in the mouse embryo and the pregnant uterus.

Neuropsin is a novel serine protease whose mRNA is expressed in the mouse central nervous system. We examined the expression of neuropsin mRNA during embryonic development using Northern and in situ hybridization in non-neural tissues. The pregnant uterus showed strong expression of neuropsin mRNA, whereas the nonpregnant uterus did not express this mRNA. Expression was first detected in the primary decidual zone at 5.5 days post coitum and was maximized at 10 days post coitum, decreasing remarkably thereafter. During mouse organogenesis, neuropsin expression was observed in the developing heart, lung, thymus, pituitary, choroid plexus, and epithelial linings of the skin, oral cavity, tongue, esophagus, and forestomach. In adult mouse organs, neuropsin mRNA was expressed in epithelial tissues covered by keratinocytes with moderate density, whereas low expression was observed in lung, thymus, and spleen. Neuropsin mRNA expression in developing organs and adult keratinocytes suggests that neuropsin is associated with extracellular matrix modifications and cell migrations.

Animals↗
PubMed1997

Crystallization and preliminary X-ray analysis of neuropsin, a serine protease expressed in the limbic system of mouse brain.

Neuropsin (M(r) 25032) is a serine protease expressed in the limbic system of mouse brain. It has been implicated in various neurological processes including formation of memory and may be important as a drug target in the treatment of epilepsy. The recombinant protein was produced using a baculovirus expression system and was purified. Two crystal forms were obtained by a hanging-drop vapor-diffusion method with polyethylene glycol. Preliminary X-ray crystallographic analysis revealed that crystal form I belongs to triclinic space group P1 with unit cell dimensions a = 97.16 A, b = 97.12 A, c = 46.75 A and alpha = 99.17 degrees, beta = 99.77 degrees, gamma = 117.35 degrees. Self-rotation function analysis of these data of form I indicates the position of a noncrystallographic threefold axis. There are six molecules in the crystallographic asymmetric unit. Crystal form II also belongs to triclinic space group P1 but has unit cell dimensions of a = 38.40 A, b = 55.16 A, c = 65.37 A and alpha = 95.38 degrees, beta = 89.98 degrees, gamma = 110.46 degrees with two molecules in the crystallographic asymmetric unit. Form II has a noncrystallographic twofold axis. Intensity data to 3.1 A resolution for form I and to 2.2 A resolution for form II have been collected.

Amino Acid Sequence↗
PubMed1996

Kindling induces neuropsin mRNA in the mouse brain.

Neuropsin mRNA expression was analyzed and mapped in the mouse brains after kindling epileptogenesis by using in situ hybridization histochemistry. Dynamic increases of the neuropsin mRNA were observed in the layer II of prelimbic, somatosensory, auditory, perirhinal, entorhinal, and piriform cortices in an activity-dependent manner, though no neuropsin gene was expressed in these areas in control mice. In addition to the confirmation of our previous studies showing increases of mRNA in the hippocampus and amygdaloid complex, there were also remarkable increases of the neuropsin mRNA in the limbic areas, such as the accessory olfactory nucleus, the medial and lateral septal nucleus, the nucleus of diagonal band, the substantia innominata and the zona incerta. The dynamic activity-dependent changes of the gene expression and the site-specificity of neuropsin localization are suggesting that this molecule is implicated in cortical- and limbic-specific neuronal reorganization.

Animals↗