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Biomedical subjects

Yasuo Ihara

Publications and source records attributed to Yasuo Ihara.

At least 19 recordsLinked to original sources
PubMed2006

Dynamin-binding protein gene on chromosome 10q is associated with late-onset Alzheimer's disease.

The apolipoprotein E (APOE) gene has been consistently shown to be a major genetic risk factor; however, all cases of Alzheimer's disease (AD) cannot be attributed to the epsilon4 variant of APOE, because about half of AD patients have the APOE-epsilon3*3 genotype. To identify an additional genetic risk factor(s), we performed large-scale single nucleotide polymorphism (SNP)-based association analysis of 1526 late-onset AD patients and 1666 control subjects in a Japanese population. We prepared two independent sets consisting of exploratory and validation samples, respectively, with only the APOE-epsilon3*3 genotype, and first carried out genotyping for the exploratory set with 1206 SNPs in the region between 60 and 107 Mb on chromosome 10q that is implicated by linkage studies as containing an AD susceptibility locus. Thirty-five SNPs that showed significant values (P<0.01) were followed-up to detect any association with the validation samples. Finally, six SNPs exhibited replicated significant associations (P=0.000035-0.00048) on meta-analysis of both sets. These SNPs were clustered in a locus spanning 220 kb at genomic position 101 Mb, and three of the six SNPs were located in the dynamin-binding protein (DNMBP) gene. Quantitative real-time RT-PCR analysis demonstrated that neuropathologically confirmed AD brains exhibit a significant reduction of DNMBP mRNA compared with age-matched ones (P<0.0169). Thus, we confirmed the association of DNMBP with AD individuals with the APOE-epsilon3*3 genotype or lacking the epsilon4 allele, and DNMBP may be one of the susceptibility genes for AD.

Aged↗
PubMed2006

Equimolar production of amyloid beta-protein and amyloid precursor protein intracellular domain from beta-carboxyl-terminal fragment by gamma-secretase.

We showed previously that cells expressing wild-type (WT) beta-amyloid precursor protein (APP) or coexpressing WTAPP and WT presenilin (PS) 1/2 produced APP intracellular domains (AICD) 49-99 and 50-99, with the latter predominating. On the other hand, the cells expressing mutant (MT) APP or coexpressing WTAPP and MTPS1/2 produced a greater proportion of AICD-(49-99) than AICD-(50-99). In addition, the expression of amyloid beta-protein (Abeta) 49 in cells resulted in predominant production of Abeta40 and that of Abeta48 leads to preferential production of Abeta42. These observations suggest that epsilon-cleavage and gamma-cleavage are interrelated. To determine the stoichiometry between Abeta and AICD, we have established a 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonic acid-solubilized gamma-secretase assay system that exhibits high specific activity. By using this assay system, we have shown that equal amounts of Abeta and AICD are produced from beta-carboxyl-terminal fragment (C99) by gamma-secretase, irrespective of WT or MTAPP and PS1/2. Although various Abeta species, including Abeta40, Abeta42, Abeta43, Abeta45, Abeta48, and Abeta49, are generated, only two species of AICD, AICD-(49-99) and AICD-(50-99), are detected. We also have found that M233T MTPS1 produced only one species of AICD, AICD-(49-99), and only one for its counterpart, Abeta48, in contrast to WT and other MTPS1s. These strongly suggest that epsilon-cleavage is the primary event, and the produced Abeta48 and Abeta49 rapidly undergo gamma-cleavage, resulting in generation of various Abeta species.

Amyloid Precursor Protein Secretases↗
PubMed2006

DAPT-induced intracellular accumulations of longer amyloid beta-proteins: further implications for the mechanism of intramembrane cleavage by gamma-secretase.

Gamma-secretase cleaves the transmembrane domain of beta-amyloid precursor protein at multiple sites. These are referred to as gamma-, zeta-, and epsilon-cleavages. We showed previously that DAPT, a potent dipeptide gamma-secretase inhibitor, caused differential accumulations of longer amyloid beta-proteins (Abetas) (Abeta43 and Abeta46) in CHO cells that are induced to express the beta C-terminal fragment (CTF). To learn more about the cleavage mechanism by gamma-secretase, CHO cell lines coexpressing betaCTF and wild-type or mutant presenilin (PS) 1/2 were generated and treated with DAPT. In all cell lines treated with DAPT, as the levels of Abeta40 decreased, Abeta46 accumulated to varying extents. In wild-type PS1 or M146L mutant PS1 cells, substantial amounts of Abeta43 and Abeta46 accumulated. In contrast, this was not the case with wild-type PS2 cells. In M233T mutant PS1 cells, significant amounts of Abeta46 and Abeta48 accumulated differentially, whereas in N141I mutant PS2 cells, large amounts of Abeta45 accumulated concomitantly with a large decrease in Abeta42 levels. Most interestingly, in G384A mutant PS1 cells, there were no significant accumulations of longer Abetas except for Abeta46. Abeta40 was very susceptible to DAPT, but other Abetas were variably resistant. Complicated suppression and accumulation patterns by DAPT may be explained by stepwise processing of betaCTF from a zeta- or epsilon-cleavage site to a gamma-cleavage site and its preferential suppression of gamma-cleavage over zeta- or epsilon-cleavage.

Amyloid Precursor Protein Secretases↗
PubMed2006

Neurofibrillary tangles/paired helical filaments (1981-83).

Neurofibrillary tangles, one of the hallmarks of Alzheimer's disease, had been a target of modern neuropathology based on electron microscopy. In 1960s their unit fibrils were found to be paired helical filaments (PHF), the unique appearance of which attracted many researchers to their nature. In the late 1970s, a keen interest in their constituents at the molecular levels had increasingly grown, but electron microscopic approach failed to address the issue. I describe here what was going on at the turning point when electron microscopic study yielded immunocytochemical approach and direct characterization by isolation, with some emphasis on the situation in Japan. Personal memories are provided about Dr Selkoe's lab (1981-1982) in Mailman Research Center, Belmont, Boston, where we encountered a series of remarkable properties of PHF. How insolubility of PHF, and smearing on the blot was found is described.

Alzheimer Disease↗
PubMed2005

Evolution of male parental care and female multiple mating: game-theoretical and two-locus diploid models.

Males gain a fitness benefit by mating with many females, whereas the number of progeny per female does not increase as a function of additional mates. Furthermore, males run the risk of investing in the offspring of other males if they provide parental care. Nevertheless, in various species, males provide parental care, and females mate with multiple males. We investigate a game-theoretical model in which females gain a direct benefit by multiple mating from the paternal care they elicit for their offspring. The parameters that directly favor male parental care, such as small cost of paternal care, have indirect positive effects on the evolution of female multiple mating, while they have negative effects in the opposite case. Both traits are more likely to evolve when the number of matings is smaller. The individual-based model of a diploid two-locus, two-allelic genetic model confirms the result.

Animals↗
PubMed2005

Blocking the cleavage at midportion between gamma- and epsilon-sites remarkably suppresses the generation of amyloid beta-protein.

To examine how gamma- and epsilon-cleavages of beta-amyloid precursor protein (APP) are related, each cleavage site was replaced with a stretch of Trp that cannot be cleaved by gamma-secretase. Replacement of the gamma- or epsilon-site significantly suppressed secretion of amyloid beta-protein (Abeta), and produced longer Abeta or longer APP intracellular domain, respectively. This cleavage at the midportion between gamma- and epsilon-sites was also gamma-secretase-dependent. Blocking this cleavage with a Trp stretch remarkably suppressed Abeta generation, indicating that the midportion cleavage is required for the generation of Abeta.

Amino Acid Sequence↗
PubMed2005

Random mutagenesis of presenilin-1 identifies novel mutants exclusively generating long amyloid beta-peptides.

Familial Alzheimer disease-causing mutations in the presenilins increase production of longer pathogenic amyloid beta-peptides (A beta(42/43)) by altering gamma-secretase activity. The mechanism underlying this effect remains unknown, although it has been proposed that heteromeric macromolecular complexes containing presenilins mediate gamma-secretase cleavage of the amyloid beta-precursor protein. Using a random mutagenesis screen of presenilin-1 (PS1) for PS1 endoproteolysis-impairing mutations, we identified five unique mutants, including R278I-PS1 and L435H-PS1, that exclusively generated a high level of A beta43, but did not support physiological PS1 endoproteolysis or A beta40 generation. These mutants did not measurably alter the molecular size or subcellular localization of PS1 complexes. Pharmacological studies indicated that the up-regulation of activity for A beta43 generation by these mutations was not further enhanced by the difluoroketone inhibitor DFK167 and was refractory to inhibition by sulindac sulfide. These results suggest that PS1 mutations can lead to a wide spectrum of changes in the activity and specificity of gamma-secretase and that the effects of PS1 mutations and gamma-secretase inhibitors on the specificity are mediated through a common mechanism.

Allosteric Site↗
PubMed2005

Longer forms of amyloid beta protein: implications for the mechanism of intramembrane cleavage by gamma-secretase.

Gamma-cleavage of beta-amyloid precursor protein (APP) in the middle of the cell membrane generates amyloid beta protein (Abeta), and epsilon-cleavage, approximately 10 residues downstream of the gamma-cleavage site, releases the APP intracellular domain (AICD). A significant link between generation of Abeta and AICD and failure to detect AICD41-99 led us to hypothesize that epsilon-cleavage generates longer Abetas, which are then processed to Abeta40/42. Using newly developed gel systems and an N-end-specific monoclonal antibody, we have identified the longer Abetas (Abeta1-43, Abeta1-45, Abeta1-46, and Abeta1-48) within the cells and in brain tissues. The production of these longer Abetas as well as Abeta40/42 is presenilin dependent and is suppressed by {1S-benzyl-4R-[1S-carbamoyl-2-phenylethylcarbamoyl-1S-3-methylbutylcarbamoyl]-2R-hydroxy-5-phenylpentyl}carbamic acid tert-butyl ester, a transition state analog inhibitor for aspartyl protease. In contrast, N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester, a potent dipeptide gamma-secretase inhibitor, builds up Abeta1-43 and Abeta1-46 intracellularly, which was also confirmed by mass spectrometry. Notably, suppression of Abeta40 appeared to lead to an increase in Abeta43, which in turn brings an increase in Abeta46, in a dose-dependent manner. We therefore propose an alpha-helical model in which longer Abeta species generated by epsilon-cleavage is cleaved at every three residues in its carboxyl portion.

Alzheimer Disease↗
PubMed2005

Progressive neurodegeneration in C. elegans model of tauopathy.

Discovery of various mutations in the tau gene among frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) families suggests gain-of-toxic function of wild-type or mutant tau as the mechanism for extensive neuronal loss. We thus generated transgenic nematode (Caenorhabditis elegans) expressing wild-type or mutant (P301L and R406W) tau in the touch (mechanosensory) neurons. Whereas the worm expressing wild-type tau showed a small decrease in the touch response across the lifespan, the worm expressing mutant tau displayed a large and progressive decrease. When the touch neurons lost their function, neuritic abnormalities were found prominent, and microtubular loss became remarkable in the later stage. A substantial fraction of degenerating neurons developed tau accumulation in the cell body and neuronal processes. This neuronal dysfunction is not related to the apoptotic process because little recovery from touch abnormality was observed in the ced-3 or ced-4-deficient background. Expression of GSK3 brought about slight deterioration in the touch response, while expression of HSP70 led to some improvement.

Animals↗
PubMed2005

Visualization of newly deposited tau in neurofibrillary tangles and neuropil threads.

Neurofibrillary tangles (NFTs) and neuropil threads (NTs), the major hallmark of Alzheimer disease (AD), are composed of the microtubule-associated protein tau that has undergone posttranslational modifications, including deamidation and isomerization on asparaginyl or aspartyl residues. Because such modifications represent protein aging, we generated 2 antibodies, TM4, specific for Asp-387 of tau, and iD387, specific for isoAsp-387 of tau, to investigate the evolution of NFTs and NTs. On Western blots of Sarkosyl-insoluble fractions, TM4 strongly labeled paired helical filament-tau (PHF-tau), whereas iD387 preferentially labeled PHF smear. Thus, it is reasonable to postulate that TM4-labeled tau (unmodified tau species) represents more recent deposition, and iD387-labeled tau (modified tau species) represents earlier deposition. Unexpectedly, TM4 immunostained even highly evolved NFTs, suggesting that deposition of newly produced tau continues until neuronal death. iD387 labeled the whole profile of NFTs up to distal dendritic branches, whereas TM4 staining was localized to particular portions of NFTs in proximal dendrites and neuronal perikarya. In NTs, TM4 preferentially labeled the outer portion, whereas iD387 intensely labeled the core portion. Based on TM4-positive NFT counts and total NFT counts, we speculate that NFTs in the human hippocampus are produced at a constant rate irrespective of the disease stage.

Alzheimer Disease↗
PubMed2005

Semaphorin3A signalling is mediated via sequential Cdk5 and GSK3beta phosphorylation of CRMP2: implication of common phosphorylating mechanism underlying axon guidance and Alzheimer's disease.

Collapsin response mediating protein-2 (CRMP2) has been identified as an intracellular protein mediating Semaphorin3A (Sema3A), a repulsive guidance molecule. In this study, we demonstrate that cyclin-dependent kinase 5 (Cdk5) and glycogen synthase kinase 3beta (GSK3beta) plays a critical role in Sema3A signalling. In In vitro kinase assay, Cdk5 phosphorylated CRMP2 at Ser522, while GSK3beta did not induce any phosphorylation of CRMP2. Phosphorylation by GSK3beta was exclusively observed in Cdk5-phosphorylated CRMP2, but barely in CRMP2T509A. These results indicate that Cdk5 primarily phosphorylates CRMP2 at Ser522 and GSK3beta secondarily phosphorylates at Thr509. The dual-phosphorylated CRMP2, but not non-phosphorylated or single-phosphorylated CRMP2, is recognized with the antibody 3F4, which is highly reactive with the neurofibrillary tangles of Alzheimer's disease. 3F4 recognized the CRMP2 in the wild-type but not cdk5-/- mouse embryonic brain lysates. The phosphorylation of CRMP2 at Ser522 caused reduction of its affinity to tubulin. In dorsal root ganglion neurones, Sema3A stimulation enhanced the levels of the phosphorylated form of CRMP2 detected by 3F4. Over-expression of CRMP2 mutant substituting either Ser522 or Thr509 to Ala attenuates Sema3A-induced growth cone collapse response. These results suggest that the sequential phosphorylation of CRMP is an important process of Sema3A signalling and the same mechanism may have some relevance to the pathological aggregation of the microtubule-associated proteins.

Alzheimer Disease↗
PubMed2005

[A proposal for diagnostic and clinical assessment criteria for Alzheimer's disease].

To clarify the risk and associated genes of Alzheimer's disease by genome-wide screening, a Japanese study group was organized in 2000 under Yasuo Ihara, Tokyo University, supported by a Grant-in-Aid for Science Research on Priority Areas (C) -Advanced Brain Science Project from Ministry of Education, Culture, Sports, Science and Technology, Japan. This is the first Japanese consortium study under permission of the ethical committees of the enrolled institutes based on the ethics guidelines for human genome/gene analysis research, Ministry of Education, Culture, Sports, Science and Technology Ministry of Health, Labor and Welfare Ministry of Economy, Trade and Industry. In this project, 2,000 genome samples from patients with Alzheimer's disease, 2,000 control subjects, and 200 siblings affected with Alzheimer's disease are collected and analyzed. For this purpose, it is necessary to analyze samples from accurately diagnosed Alzheimer patients and controls using standard criteria for diagnosis and neuropsychological evaluation, which have been confirmed by an evidence-based studying a Japanese population. Here, we propose criteria for the diagnosis and clinical assessment of Alzheimer's disease. This proposal consists of a definition of Alzheimer's disease based on recent advances in research, diagnostic criteria based on DSM-IV, NINCDS-ADRDA and ICD-10, exclusion criteria for other dementia disorders, routine and detailed tests for neuropsychological and laboratory evaluations, criteria for neuroimaging and biomarkers, definitive diagnostic criteria and classification of clinical subtypes.

Alzheimer Disease↗
PubMed2004

Truncated carboxyl-terminal fragments of beta-amyloid precursor protein are processed to amyloid beta-proteins 40 and 42.

We previously showed that beta-amyloid precursor protein (APP) is cleaved not only in the middle of the membrane (gamma-cleavage) but also at novel cleavage sites close to the membrane/cytoplasmic boundary (epsilon-cleavage), releasing APP intracellular domains (AICDs) 49-99 and 50-99. To learn more about the relationship between gamma- and epsilon-cleavage, C-terminally truncated carboxyl-terminal fragments (CTFs) of APP, especially CTFs1-48 and 1-49 (the postulated products that are generated by epsilon-cleavage), were transiently expressed in CHO cells. Most importantly, the cells expressing CTF1-49 secreted predominantly amyloid beta-protein (Abeta) 40, while those expressing CTF1-48 secreted preferentially Abeta42. This supports our assumption that epsilon-cleavage precedes Alphabeta production and that preceding epsilon-cleavage determines the preference for the final Abeta species. The gamma-secretase inhibitors, L-685,458 and DAPT, suppressed Abeta production from CTF1-49. Regarding Abeta production from CTF1-48, L-685,458 suppressed it, but DAPT failed to do so. A dominant negative mutant of presenilin 1 suppressed the production of Abeta40 and 42 from both CTFs1-48 and 1-49. These data should shed significant light into the mechanism of Abeta production.

Amino Acid Sequence↗
PubMed2004

Gem GTPase and tau: morphological changes induced by gem GTPase in cho cells are antagonized by tau.

A series of observations have indicated that tau, one of the major microtubule-associated proteins, is involved in neuronal cell morphogenesis and axonal maintenance. Tau is also the major component of paired helical filaments found in brains affected by Alzheimer's disease. To explore an as yet unidentified role of tau in vivo, approximately 11,000 mRNAs were profiled from tau-deficient mouse brains and compared with those from control brains at the same ages. The expression of Gem GTPase, a small GTP-binding protein of the ras superfamily, was significantly increased in the brains of tau-deficient mice at 8 weeks of age. Because Gem GTPase is a negative regulator of the Rho-Rho kinase pathway for cytoskeletal organization, this protein was transiently overexpressed in Chinese hamster ovary cells that do not express tau. Overexpression of Gem GTPase induced a marked elongation of Chinese hamster ovary cells, and simultaneous expression of tau eliminated this effect, although tau did not bind directly to Gem GTPase. This anti-elongation activity of tau was attributed to its microtubule-binding domain, and homologous domains of microtubule-associated proteins 2 and 4 exhibited similar antagonistic activities. Taken together, the present results indicate that the level of Gem GTPase and its cell elongation activity are modulated by tau and suggest that tau may be involved in a Gem GTPase-mediated signal transduction pathway.

Amino Acid Sequence↗
PubMed2004

Glypican-1 as an Abeta binding HSPG in the human brain: its localization in DIG domains and possible roles in the pathogenesis of Alzheimer's disease.

Previous studies have suggested that heparan sulfate proteoglycans (HSPGs) play a role in deposition of beta-amyloid protein (Abeta) in the Alzheimer's disease (AD) brain. In the present study, we demonstrated that glypican-1 can bind fibrillar Abeta, and the binding is mainly mediated by heparan sulfate (HS) chains. Further analysis revealed that glypican-1 is the major HSPG localized in detergent-insoluble glycosphingolipid-enriched (DIG) domains where all machineries for Abeta production exist and Abeta is accumulated as monomeric and oligomeric forms. Immunohistochemical studies demonstrated that glypican-1 is localized in primitive plaques as well as classic plaques. Moreover, overexpression of glypican-1 and amyloid precursor protein in SH-SY5Y cells resulted in reduced cell viability and made cells more susceptible to thapsigargin-induced stress and Abeta toxicity. The results raise the possibility that glypican-1 interacts with oligomerized or polymerized Abeta in such a specific compartment as DIG, resulting not only in amyloid deposition in senile plaques of AD brain, but also in accelerating neuronal cell death in response to stress and Abeta.

Aged↗
PubMed2004

Cultural niche construction and the evolution of small family size.

A model of cultural niche construction with two culturally transmitted traits is examined. The frequency of individuals with a certain general predisposition, which is transmitted vertically, plays a role as the cultural background, or the cultural niche, of the population. The cultural background determines the rate of oblique, relative to vertical, transmission of another cultural trait that affects fertility of individuals. It is assumed that individuals with fewer offspring are more likely to achieve social roles that influence the succeeding generation and are therefore overrepresented as transmitters in the process of oblique transmission. Our model suggests that even a slight overrepresentation of those with fewer offspring can drive the evolution of small family size, provided that the rate of oblique transmission depends strongly on the cultural background. In addition, our model may help to explain the time lag between the decrease in death rates and the subsequent decrease in birth rates during the demographic transition of industrializing societies.

Cultural Characteristics↗
PubMed2004

Molecular aging of tau: disulfide-independent aggregation and non-enzymatic degradation in vitro and in vivo.

Smearing from high-molecular-mass regions to low-molecular-mass regions on western blot is the most striking observation of the tau making up paired helical filaments in brain tissues affected by Alzheimer's disease. Because our previous study showed site-specific deamidation/isomerization in the smeared tau in vivo, a feature of protein aging, recombinant tau was subjected to prolonged (up to 90 days) in vitro incubation. Carboxymethylated tau at approximately 50 kDa gradually disappeared and was converted to dimers and to high- and low-molecular-mass smearing. In addition, the same site-specific deamidation/isomerization as previously identified in the smeared tau in vivo emerged. Most importantly, tau was spontaneously degraded, generating fragments that start from bulky residues next to asparaginyl residues. This spontaneous degradation of tau probably represents non-enzymatic cleavage through the formation of succinimide intermediates. Similar degradation products starting from the bulky residues next to asparaginyl residues were found in the smeared tau in vivo partially purified from the homogenates from Alzheimer's disease brains.

Aging↗