Biomedical subjects

Y Nagamine

Publications and source records attributed to Y Nagamine.

At least 19 recordsLinked to original sources

PubMed2004

Non-transcallosal ipsilateral area 3b responses to median nerve stimulus.

We report two patients with left hemisphere lesions who had no normal left hemispheric responses to right median nerve stimulus on magnetoencephalography but displayed right area 3b responses. One patient had suffered a severe left hemispheric contusion and the other left hemispheric infarction. Equivalent current dipoles of these ipsilateral responses were detected on the central sulcus adjacent to the location of the N20m response to left median nerve stimulus. The somatosensory afferent pathway from the hand may extend directly to the ipsilateral area 3b without following the transcallosal pathway in at least part of the population.

Adult↗

PubMed2004

Genetic mapping of quantitative trait loci affecting body weight, egg character and egg production in F2 intercross chickens.

Phenotypic measurements of chicken egg character and production traits are restricted to mature females only. Marker assisted selection of immature chickens using quantitative trait loci (QTL) has the potential to accelerate the genetic improvement of these traits in the chicken population. The QTL for 12 traits (i.e. body weight (BW), six for egg character, three for egg shell colour and two for egg production) of chickens were identified. An F2 population comprising 265 female chickens obtained by crossing White Leghorn and Rhode Island Red breeds and genotyped for 123 microsatellite markers was used for detecting QTL. Ninety-six markers were mapped on 25 autosomal linkage groups, and 13 markers were mapped on one Z chromosomal linkage group. Eight previous unmapped markers were assigned to their respective chromosomes in this study. Significant QTL were detected for BW on chromosomes 4 and 27, egg weight on chromosome 4, the short length of egg on chromosome 4, and redness of egg shell colour (using the L*a*b* colour system) on chromosome 11. A significant QTL on the Z chromosome was linked with age at first egg. Significant QTL could account for 6-19% of the phenotypic variance in the F2 population.

Analysis of Variance↗

PubMed2002

Simple deterministic identity-by-descent coefficients and estimation of QTL allelic effects in full and half sibs.

Accurate and rapid methods for the detection of quantitative trait loci (QTLs) and evaluation of consequent allelic effects are required to implement marker-assisted selection in outbred populations. In this study, we present a simple deterministic method for estimating identity-by-descent (IBD) coefficients in full- and half-sib families that can be used for the detection of QTLs via a variance-component approach. In a simulated dataset, IBD coefficients among sibs estimated by the simple deterministic and Markov chain Monte Carlo (MCMC) methods with three or four alleles at each marker locus exhibited a correlation of greater than 0.99. This high correlation was also found in QTL analyses of data from an outbred pig population. Variance component analysis used both the simple deterministic and MCMC methods to estimate IBD coefficients. Both procedures detected a QTL at the same position and gave similar test statistics and heritabilities. The MCMC method, however, required much longer computation than the simple method. The conversion of estimated QTL genotypic effects into allelic effects for use in marker-assisted selection is also demonstrated.

Animals↗

PubMed2001

p53 Phosphorylation at serine 15 is required for transcriptional induction of the plasminogen activator inhibitor-1 (PAI-1) gene by the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine.

The alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) is a widely spread environmental carcinogen that causes DNA lesions leading to cell killing. MNNG can also induce a cell-protective response by inducing the expression of DNA repair/transcription-related genes. We recently demonstrated that urokinase-type plasminogen activator, an extracellular protease to which no DNA repair functions have been assigned, was induced by MNNG. Here, we show that the physiological inhibitor of urokinase-type plasminogen activator, PAI-1, is also induced by MNNG in a p53-dependent fashion, because MNNG induced PAI-1 in p53-expressing cells but not in p53-/- cells. MNNG induced p53 phosphorylation at serine 15, resulting in stabilization of the p53 protein, and this phosphorylation event was central for p53-dependent PAI-1 transcription. Finally, we showed that PAI-1 transcriptional induction by MNNG required a p53-responsive element located at -136 base pairs in the PAI-1 promoter, because specific mutation of this site abrogated the induction. Because PAI-1 is a prognostic factor in many metastatic cancers, being involved in the control of tumor invasiveness, our finding that a genotoxic agent induces the PAI-1 gene via p53 adds a new feature to the role of the tumor-suppressor p53 protein. Our results also suggest the possibility that genotoxic agents contribute to tumor metastasis by inducing PAI-1 without involving genetic modification.

3T3 Cells↗

PubMed2001

Somatosensory evoked fields in comatose survivors after severe traumatic brain injury.

OBJECTIVE: To evaluate the cortical function quantitatively in patients in the chronic phase of severe traumatic brain injury. METHODS: Thirteen patients with severe traumatic brain injury due to traffic accident followed by persistent consciousness disturbance and disability were studied. Somatosensory evoked magnetic fields (SEFs) for unilateral median nerve stimulation were measured using a whole-head magnetoencephalography system. The latency and electrical current dipole (ECD) moment for the N20m, P30m, N45m and P60m components were calculated and compared with those of 14 age-matched healthy adults. RESULTS: The peak latency of N20m was longer (P<0.05) and those of P30m and N45m were shorter (P<0.01) in the patients than in normal adults. The ECD moment of N20m and P30m was smaller and that of N45m and P60m was larger in the patients than in normal adults (P<0.01). CONCLUSIONS: These results can be explained by the hypothesis that diffuse brain injury induces decreased and delayed input of the somatosensory afferent and compensational amplification of the response in the primary somatosensory cortex. Middle-latency SEFs may be applicable as a cortical functional measure for patients with severe traumatic brain injury.

Adult↗

PubMed2001

Using the mixed model for interval mapping of quantitative trait loci in outbred line crosses.

Interval mapping by simple regression is a powerful method for the detection of quantitative trait loci (QTLs) in line crosses such as F2 populations. Due to the ease of computation of the regression approach, relatively complex models with multiple fixed effects, interactions between QTLs or between QTLs and fixed effects can easily be accommodated. However, polygenic effects, which are not targeted in QTL analysis, cannot be treated as random effects in a least squares analysis. In a cross between true inbred lines this is of no consequence, as the polygenic effect contributes just to the residual variance. In a cross between outbred lines, however, if a trait has high polygenic heritability, the additive polygenic effect has a large influence on variation in the population. Here we extend the fixed model for the regression interval mapping method to a mixed model using an animal model. This makes it possible to use not only the observations from progeny (e.g. F2), but also those from the parents (F1) to evaluate QTLs and polygenic effects. We show how the animal model using parental observations can be applied to an outbred cross and so increase the power and accuracy of QTL analysis. Three estimation methods, i.e. regression and an animal model either with or without parental observations, are applied to simulated data. The animal model using parental observations is shown to have advantages in estimating QTL position and additive genotypic value, especially when the polygenic heritability is large and the number of progeny per parent is small.

Animals↗

PubMed2001

E2F1-mediated transcriptional inhibition of the plasminogen activator inhibitor type 1 gene.

Gene expression of the plasminogen activation system is cell-cycle dependent. Previously, we showed that ectopic expression of E2F1 repressed the plasminogen activator inhibitor type 1 (PAI-1) promoter in a manner dependent on the presence of DNA-binding and transactivation domains of E2F1 but independent of binding to pocket-binding proteins, suggesting a novel mechanism for E2F-mediated negative gene regulation [Koziczak, M., Krek, W. & Nagamine, Y. (2000) Mol. Cell. Biol. 20, 2014-2022]. However, it remains to be seen whether endogenous E2F can exert a similar effect. We report here that down-regulation of PAI-1 gene expression correlates with an increase in endogenous E2F activity. When cells were treated with a cdk2/4-specific inhibitor, which maintains E2F in an inactive state, the decline of serum-induced PAI-1 mRNA levels was suppressed. In mutant U2OS cells expressing a temperature-sensitive retinoblastoma protein (pRB), a shift to a permissive temperature induced PAI-1 mRNA expression. In U2OS cells stably expressing an E2F1-estrogen receptor chimeric protein that could be activated by tamoxifen, PAI-1 gene transcription was markedly reduced by tamoxifen even in the presence of cycloheximide. These results all indicate that endogenous E2F can directly repress the PAI-1 gene. DNase I hypersensitive-site analysis of the PAI-1 promoter suggested the involvement of conformation changes in chromatin structure of the PAI-1 promoter. 5' deletion analysis of the PAI-1 promoter showed that multiple sites were responsible for the E2F negative regulation, some of which were promoter dependent. Interestingly, one of these sites is a p53-binding element.

Animals↗

PubMed2001

Chicken microsatellite primers are not efficient markers for Japanese quail.

Domestic fowl or chicken (Gallus gallus) and Japanese quail (Coturnix japonica) belong to the family Phasianidae. The exchange of marker information between chicken and quail is an important step towards the construction of a high-resolution comparative genetic map in Phasianidae, which includes several poultry species of agricultural importance. We tested chicken microsatellite markers to see if they would be suitable as genetic linkage markers in Japanese quail. Twenty-six per cent (31/120) of chicken primers amplified individual loci in Japanese quail and 65% (20/31) of the amplified loci were found to be polymorphic. Eleven of the polymorphic loci were excluded as uninformative because of the lack of amplification in some individuals or high frequency of nonspecific amplification. The sequence information of the remaining nine loci revealed six of them to contain microsatellites that were nearly identical with those of the orthologous regions in chicken. For these six loci, allele frequencies were estimated in 50 unrelated quails. Although the very few chicken markers that do work well in quail could be used as anchor points for a comparative mapping, most chicken markers are not useful for studies in quail. Therefore, more effort should be committed to developing quail-specific markers rather than attempting to adapt chicken markers for work in quail.

Animals↗

PubMed2000

Pocket protein-independent repression of urokinase-type plasminogen activator and plasminogen activator inhibitor 1 gene expression by E2F1.

Expression of genes of the plasminogen activator (PA) system declines at the G(0)/G(1)-S-phase boundary of the cell cycle. We found that overexpression of E2F1-3, which acts mainly in late G(1), inhibits promoter activity and endogenous expression of the urokinase-type PA (uPA) and PA inhibitor 1 (PAI-1) genes. This effect is dose dependent and conserved in evolution. Mutation analysis indicated that both the DNA-binding and transactivation domains of E2F1 are necessary for this regulation. Interestingly, an E2F1 mutant lacking the pRB-binding region strongly repressed the uPA and PAI-1 promoters. An E2F-mediated negative effect was also observed in pRB and p107/p130 knockout cell lines. This is the first report that E2F can act as a repressor independently of pocket proteins. Mutation of AP-1 elements in the uPA promoter abrogated E2F-mediated transcriptional inhibition, suggesting the involvement of AP-1 in this regulation. Results shown here identify E2F as an important component of transcriptional control of the PA system and thus provide new insights into mechanisms of cellular proliferation.

Animals↗

PubMed1999

Regulation by p38 mitogen-activated protein kinase of adenylate- and uridylate-rich element-mediated urokinase-type plasminogen activator (uPA) messenger RNA stability and uPA-dependent in vitro cell invasion.

MDA-MB-231 cells are highly metastatic breast tumor cells. Their high invasiveness is thought to be due to constitutively high levels of urokinase-type plasminogen activator (uPA) and its receptor. Previously (R. Nanbu et al., C. Eur. J. Biochem., 247: 169-174, 1997), we showed that uPA mRNA in these cells is stable and that mRNA degradation mediated by an AU-rich element (ARE) is impaired. Here we report that treatment of MDA-MB-231 cells with SB203580, an inhibitor of the stress-activated p38 mitogen-activated protein (MAP) kinase, strongly destabilized uPA mRNA in an ARE-dependent manner. In contrast, in LLC-PK1 and HeLa cells, uPA mRNA is unstable, and an ARE present in the 3' untranslated region plays a role in its degradation. Enhanced ARE-mediated mRNA destabilization induced by SB203580 was also observed in both LLC-PK1 and HeLa cells with a globin chimeric mRNA harboring two copies of the ARE (globin-2ARE) from uPA mRNA. Overexpression of constitutively active MKK6, a p38 upstream activator kinase, increased the stability of the globin-2ARE message in LLC-PK1 cells, confirming the participation of p38 in the regulation of ARE-mediated mRNA decay. Interestingly, the half-life of the uPA mRNA in the three cell lines studied correlated with the basal levels of active p38. SB203580 treatment of MDA-MB-231 cells decreased cell-associated uPA activity and dramatically reduced in vitro cell invasiveness. These results suggest the participation of p38 in the control of invasiveness through regulation of the stability of uPA and uPA receptor mRNA, which is also destabilized by p38.

3' Untranslated Regions↗

PubMed1999

The plasminogen activator system: biology and regulation.

The regulation of plasminogen activation involves genes for two plasminogen activators (tissue type and urokinase type), two specific inhibitors (type 1 and type 2), and a membrane-anchored urokinase-type plasminogen-activator-specific receptor. This system plays an important role in various biological processes involving extracellular proteolysis. Recent studies have revealed that the system, through interplay with integrins and the extracellular matrix protein vitronectin, is also involved in the regulation of cell migration and proliferation in a manner independent of proteolytic activity. The genes are expressed in many different cell types and their expression is under the control of diverse extracellular signals. Gene expression reflects the levels of the corresponding mRNA, which should be the net result of synthesis and degradation. Thus, modulation of mRNA stability is an important factor in overall regulation. This review summarizes current understanding of the biology and regulation of genes involved in plasminogen activation at different levels.

Animals↗

PubMed1999

Cytoskeletal reorganization leads to induction of the urokinase-type plasminogen activator gene by activating FAK and Src and subsequently the Ras/Erk signaling pathway.

Previously, we showed that cytoskeletal reorganization (CSR) induced by colchicine or cyochalasins leads to activation of the urokinase-type plasminogen activator (uPA) gene in LLC-PK(1) cells via the Ras/Erk signaling pathway [Irigoyen et al. (1997) J. Biol. Chem. 272, 1904]. It remained to be seen how CSR activates Ras/Erk signaling. Changes in cell morphology triggered by extracellular signals are often mediated by integrin-associated proteins, such as focal adhesion kinase (FAK) and Src. We found that CSR induced the activation of FAK and Src and the association of FAK and Shc, a signaling molecule linking growth factor receptor tyrosine kinase and Grb2. Furthermore, expression of either FRNK, a kinase-minus FAK-like molecule acting as a dominant negative FAK, or a dominant negative Src suppressed CSR-induced uPA gene promoter activation. These results suggest that cells respond to a morphology change, using the cytoskeleton as a sensor, by activating FAK and Src and subsequently the Ras/Erk signaling pathway.

Animals↗

PubMed1999

A mammalian ortholog of Drosophila timeless, highly expressed in SCN and retina, forms a complex with mPER1.

BACKGROUND: It is now becoming clear that the circadian rhythm of behaviours and hormones arises from a rhythm at the level of gene expression, and that mammals and Drosophila essentially use homologous genes as molecular gears in the control of circadian oscillation. In Drosophila, the period and timeless genes form a functional unit of the clock and its autoregulatory feedback loop for circadian rhythm. However, in mammals, the counterpart of timeless has not been found. RESULTS: We have isolated a mammalian homologue of timeless, mTim, from the mouse brain. mTim is highly expressed, with a weak or absent rhythm in the suprachiasmatic nucleus, the mammalian circadian centre. In the retina, mTim mRNA was found to be expressed with a circadian rhythm, and a particularly robust cycle was observed in the presence of light/dark cycles. We demonstrated that mTIM physically associates with mPER1 in vitro and in the nuclei of cultured COS7 cells. CONCLUSIONS: We have reported the isolation of the mouse timeless cDNA, the expression of the mTim mRNA and an interaction of mTIM with mPER1. These results indicate that the autoregulatory feedback mechanism of circadian oscillation of the period gene may also be conserved in mammals.

Amino Acid Sequence↗

PubMed1999

Detection of human papilloma virus type 56 in extragenital Bowen's disease.

A case of Bowen's disease arising on the medial part of the first metatarsal bone of an 81-year-old Japanese woman is described. Histopathologically, proliferation of atypical cells was found throughout the epidermis. Electronmicroscopy revealed virus particles 40-50 nm in diameter in the nuclei of tumour cells at the granular cells just on or below the horny layer. Positive bands were obtained by polymerase chain reaction using a consensus primer of human papilloma virus L1 portion. Sequencing analysis of the amplified DNA revealed the same base sequences and homology as human papilloma virus 56. To the best of our knowledge, this case is the first report in which human papilloma virus 56 was found in a case of extragenital Bowen's disease. We consider it important to understand that human papilloma virus 56, often found in cervical lesions, can be detected in extragenital Bowen's diseases.

Aged↗

PubMed1999

Mode of action of anhydrofulvic acid against Candida utilis ATCC 42402 under acidic condition.

The mode of action of anhydrofulvic acid against Candida utilis ATCC 42402 was investigated under acidic conditions. Anhydrofulvic acid inhibited the incorporation of radioactive precursors into DNA, RNA, protein and lipid fractions. Although it did not induce leakage of intracellular materials from the treated cells, it had inhibitory effects on both endogenous and exogenous cellular respiration. Moreover, it inhibited mitochondrial respiration of Candida utilis ATCC 42402 using both succinate and cytochrome c as respiratory substrates, but not using NADH. Unexpectedly, the inhibition against isolated mitochondria was observed at pH 7.0. These results suggested that the action site against the respiratory inhibition of anhydrofulvic acid might be involved in succinate dehydrogenase, complex II in the mitochondrial electron transport chain of the yeast cells. Judging from the inhibitory effect of anhydrofulvic acid on mitochondria detected at pH 7.0, it was postulated that the antifungal activity at a low pH level might depend on the elevation of drug permeability to the cell membrane under acidic conditions.

Antifungal Agents↗

PubMed1999

Transcriptional regulation of the murine urokinase-type plasminogen activator gene in skeletal myoblasts.

We have previously shown that urokinase-type plasminogen activator (uPA) is highly expressed in murine C2C12 myoblasts and that antibodies against uPA are able to block both myoblast fusion and differentiation. Here we show the characterization of cis-acting elements in the mouse uPA promoter in vitro which are involved in uPA gene expression in C2C 12 myoblast cells. DNase I hypersensitive (HS) site analysis revealed the presence of three HS sites in myoblasts. Deletion analysis of stably transfected uPA-promoter constructs revealed that at least two of the three HS sites accounted for the high transcriptional expression in C2C12 cells. One was located at -2.4 kb and corresponded to a known PEA3/AP1A element and the other one was located at -4.9 kb and contained a CArG box and a CRE element. So far, no regulatory function had been assigned to this CRE/CArG element. Both HS sites alone were able to activate transcription of a heterologous promoter and showed a cooperative effect when placed together. Electrophoretic mobility-shift assays using myoblast nuclear extracts and specific antibodies demonstrated that cJun, JunD and ATF2 bound to the PEA3/AP1A element, whereas the CRE/CArG element bound SRF. Altogether, these results suggest that high uPA expression in myoblasts is dependent on the cooperation of two regulatory sites in the uPA promoter.

Animals↗

PubMed1998

Factors that cause the beta-anomeric preference of Na+/glucose cotransporter for intestinal transport of monosaccharide conjugates.

The intestinal transport of glucose- and galactose-conjugated acetaminophen (APAP glycoside) by Na+/glucose cotransporter (SGLT1) was studied. SGLT1-mediated transport of APAP glycosides preferred glucoside>galactoside and beta-anomer>alpha-anomer. These preferences agree with previous studies. NMR spectroscopic and molecular modeling studies indicated that the conformation of the glucose ring of alpha- and beta-glucosides of APAP, as well as glycosides in previous studies, is in the 4C1 chair form, the same form as glucose itself. Molecular dynamics analysis also indicated that the glucose ring was in the 4C1 chair form, and that there are differences between the rotational spaces of aglycones and hydroxy groups of glucose moieties between anomers. Therefore, we conclude that the beta-anomeric preference of glucose conjugate transport by SGLT1 is not due to the conformation of the glucose ring, but to the configuration of the aglycone at C-1 of the monosaccharide moiety.

Acetaminophen↗

PubMed1998

Interaction of the DNA modifying proteins VirD1 and VirD2 of Agrobacterium tumefaciens: analysis by subcellular localization in mammalian cells.

Interaction between Agrobacterium tumefaciens and plants provides a unique example of interkingdom gene transfer. Agrobacterium, a plant pathogen, is capable to stably transform the plant cell with a segment of its own DNA called T-DNA (transferred DNA). This process depends, among others, on the specialized bacterial virulence proteins VirD1 and VirD2 that excise the T-DNA from its adjacent sequences. Subsequent to transfer to the plant cell, the virulence protein VirD2, through its nuclear localization signal (NLS), is believed to guide the T-DNA to the nucleus. The T-DNA then is integrated into the plant genome. Although both of these proteins are essential for bacterial virulence, physical interaction of them has not been analyzed so far. We studied associations between these proteins by expressing them in mammalian cells and by testing for intracellular localization and colocalization. When expressed in human cells [HeLa, human embryo kidney (HEK) 293], the VirD2 protein homogeneously distributed over the nucleoplasm. The presence of any of two NLSs, on the N and C termini of VirD2, was sufficient for its efficient nuclear localization whereas deletion of both NLSs rendered the protein cytoplasmic. However, this double NLS mutant was translocated to the nucleus in the presence of wild-type VirD2 protein, implying VirD2-VirD2 interaction. The VirD1 protein, by itself localized in the cytoplasm, moved to the nucleus when coexpressed with the VirD2 protein, suggesting VirD1-VirD2 interaction. This interaction was confirmed by coimmunoprecipitation tests. Of interest, both proteins coimported to the nucleus showed a similar, peculiar sublocalization. The data are discussed in terms of functions of the VirD proteins. In addition, coimport of proteins into nuclei is suggested as a useful system in studying individual protein-protein interactions.

Agrobacterium tumefaciens↗