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61.
Sakai H Suzuki S Mizuguchi T Imoto K Yamashita Y Doi H Kikuchi M Tsurusaki Y Saitsu H Miyake N Masuda M Matsumoto N 《Human genetics》2012,131(4):591-599
Aortic aneurysm and/or dissection (AAD) is a life-threatening condition, and several syndromes are known to be related to AAD. In this study, two new technologies, resequencing array technology (ResAT) and next-generation sequencing (NGS), were used to analyze eight genes associated with syndromic AAD in 70 patients with non-syndromic AAD. Eighteen sequence variants were detected using both ResAT and NGS. In addition one of these sequence variants was detected by ResAT only and two additional variants by NGS only. Three of the 18 variants are likely to be pathogenic (in 4.3% of AAD patients and in 8.6% of a subset of patients with thoracic AAD), highlighting the importance of genetic analysis in non-syndromic AAD. ResAT and NGS similarly detected most, but not all, of the variants. Resequencing array technology was a rapid and efficient method for detecting most nucleotide substitutions, but was unable to detect short insertions/deletions, and it is impractical to update custom arrays frequently. Next-generation sequencing was able to detect almost all types of mutation, but requires improved informatics methods. 相似文献
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Hirotomo Takatsuka Ryoko Ohno Masaaki Umeda 《The Plant journal : for cell and molecular biology》2009,59(3):475-487
Cyclin-dependent kinases (CDKs) play an essential role in cell cycle regulation during the embryonic and post-embryonic development of various organisms. Full activation of CDKs requires not only binding to cyclins but also phosphorylation of the T-loop domain. This phosphorylation is catalysed by CDK-activating kinases (CAKs). Plants have two distinct types of CAKs, namely CDKD and CDKF; in Arabidopsis, CDKF;1 exhibits the highest CDK kinase activity in vitro . We have previously shown that CDKF;1 also functions in the activation of CDKD;2 and CDKD;3 by T-loop phosphorylation. Here, we isolated the knockout mutants of CDKF;1 and showed that they had severe defects in cell division, cell elongation and endoreduplication. No defect was observed during embryogenesis, suggesting that CDKF;1 function is primarily required for post-embryonic development. In the cdkf;1 mutants, T-loop phosphorylation of CDKA;1, an orthologue of yeast Cdc2/Cdc28p, was comparable to that in wild-type plants, and its kinase activity did not decrease. In contrast, the protein level and kinase activity of CDKD;2 were significantly reduced in the mutants. Substitution of threonine-168 with a non-phosphorylatable alanine residue made CDKD;2 unstable in Arabidopsis tissues. These results indicate that CDKF;1 is dispensable for CDKA;1 activation but is essential for maintaining a steady-state level of CDKD;2, thereby suggesting the quantitative regulation of a vertebrate-type CAK in a plant-specific manner. 相似文献
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Minoru Kishi Yasuhisa Abe Hirotomo Sasaki Mami Shimizu Takashi Arai Yasuyo Okumachi Hiroaki Moriyama Kenta Hara Koichi Yokono Masao Nagata 《Biochemical and biophysical research communications》2010,394(1):228-232
Antigen-specific regulatory CD4+ T cells have been described but there are few reports on regulatory CD8+ T cells. We generated islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)-specific regulatory CD8+ T cells from 8.3-NOD transgenic mice. CD8+ T cells from 8.3-NOD splenocytes were cultured with IGRP, splenic dendritic cells (SpDCs), TGF-β, and all-trans retinoic acid (ATRA) for 5 days. CD8+ T cells cultured with either IGRP alone or IGRP and SpDCs in the absence of TGF-β and ATRA had low Foxp3+ expression (1.7 ± 0.9% and 3.2 ± 4.5%, respectively). In contrast, CD8+ T cells induced by exposure to IGRP, SpDCs, TGF-β, and ATRA showed the highest expression of Foxp3+ in IGRP-reactive CD8+ T cells (36.1 ± 10.6%), which was approximately 40-fold increase compared with that before induction culture. CD25 expression on CD8+ T cells cultured with IGRP, SpDCs, TGF-β, and ATRA was only 7.42%, whereas CD103 expression was greater than 90%. These CD8+ T cells suppressed the proliferation of diabetogenic CD8+ T cells from 8.3-NOD splenocytes in vitro and completely prevented diabetes onset in NOD-scid mice in cotransfer experiments with diabetogenic splenocytes from NOD mice in vivo. Here we show that exposure to ATRA and TGF-β induces CD8+Foxp3+ T cells ex vivo, which suppress diabetogenic T cells in vitro and in vivo. 相似文献
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Hirotomo Saitsu Jun Tohyama Kiyoshi Egawa Ippei Okada Hitoshi Osaka Tomonori Furukawa Hideki Hoshino Yasuo Hachiya Shinji Saitoh Kiyomi Nishiyama Noriko Miyake Kenji Hayashi Kazuhiro Ogata Atsuo Fukuda 《American journal of human genetics》2010,86(6):881-891
A de novo 9q33.3-q34.11 microdeletion involving STXBP1 has been found in one of four individuals (group A) with early-onset West syndrome, severe hypomyelination, poor visual attention, and developmental delay. Although haploinsufficiency of STXBP1 was involved in early infantile epileptic encephalopathy in a previous different cohort study (group B), no mutations of STXBP1 were found in two of the remaining three subjects of group A (one was unavailable). We assumed that another gene within the deletion might contribute to the phenotype of group A. SPTAN1 encoding α-II spectrin, which is essential for proper myelination in zebrafish, turned out to be deleted. In two subjects, an in-frame 3 bp deletion and a 6 bp duplication in SPTAN1 were found at the initial nucleation site of the α/β spectrin heterodimer. SPTAN1 was further screened in six unrelated individuals with WS and hypomyelination, but no mutations were found. Recombinant mutant (mut) and wild-type (WT) α-II spectrin could assemble heterodimers with β-II spectrin, but α-II (mut)/β-II spectrin heterodimers were thermolabile compared with the α-II (WT)/β-II heterodimers. Transient expression in mouse cortical neurons revealed aggregation of α-II (mut)/β-II and α-II (mut)/β-III spectrin heterodimers, which was also observed in lymphoblastoid cells from two subjects with in-frame mutations. Clustering of ankyrinG and voltage-gated sodium channels at axon initial segment (AIS) was disturbed in relation to the aggregates, together with an elevated action potential threshold. These findings suggest that pathological aggregation of α/β spectrin heterodimers and abnormal AIS integrity resulting from SPTAN1 mutations were involved in pathogenesis of infantile epilepsy. 相似文献
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Oral Administration of Human T-Cell Leukemia Virus Type 1 Induces Immune Unresponsiveness with Persistent Infection in Adult Rats 总被引:3,自引:2,他引:1 下载免费PDF全文
Hirotomo Kato Yoshihiro Koya Takashi Ohashi Shino Hanabuchi Fumiyo Takemura Masahiro Fujii Hajime Tsujimoto Atsuhiko Hasegawa Mari Kannagi 《Journal of virology》1998,72(9):7289-7293
The major route of human T-cell leukemia virus type 1 (HTLV-1) infection is mother-to-child transmission caused by breast-feeding. We investigated the host immune responses to orally established persistent HTLV-1 infection in adult rats. HTLV-1-producing MT-2 cells were inoculated into immunocompetent adult rats either orally, intravenously, or intraperitoneally. HTLV-1 proviruses were detected in the peripheral blood and several organs for at least 12 weeks. Transmission of HTLV-1 to these animals was confirmed by analysis of HTLV-1 flanking regions. Despite persistent HTLV-1 presence, none of the orally inoculated rats produced detectable levels of anti-HTLV-1 antibodies, whereas all intravenously or intraperitoneally inoculated rats showed significant anti-HTLV-1 antibody responses. T-cell proliferative responses against HTLV-1 were also absent in orally inoculated rats. Our findings suggest that gastrointestinal exposure of adult rats to HTLV-1-infected cells induces persistent HTLV-1 infection in the absence of both humoral and cellular immune responses against HTLV-1. This immune unresponsiveness at primary infection may subsequently affect the host defense ability against HTLV-1. 相似文献
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Maeda K Das D Yin PD Tsuchiya K Ogata-Aoki H Nakata H Norman RB Hackney LA Takaoka Y Mitsuya H 《Journal of molecular biology》2008,381(4):956-974
C-C chemokine receptor 5 (CCR5), a member of G-protein-coupled receptors, serves as a coreceptor for human immunodeficiency virus type 1 (HIV-1). In the present study, we examined the interactions between CCR5 and novel CCR5 inhibitors containing the spirodiketopiperazine scaffolds AK530 and AK317, both of which were lodged in the hydrophobic cavity located between the upper transmembrane domain and the second extracellular loop (ECL2) of CCR5. Although substantial differences existed between the two inhibitors—AK530 had 10-fold-greater CCR5-binding affinity (Kd = 1.4 nM) than AK317 (16.7 nM)—their antiviral potencies were virtually identical (IC50 = 2.1 nM and 1.5 nM, respectively). Molecular dynamics simulations for unbound CCR5 showed hydrogen bond interactions among transmembrane residues Y108, E283, and Y251, which were crucial for HIV-1-gp120/sCD4 complex binding and HIV-1 fusion. Indeed, AK530 and AK317, when bound to CCR5, disrupted these interhelix hydrogen bond interactions, a salient molecular mechanism enabling allosteric inhibition. Mutagenesis and structural analysis showed that ECL2 consists of a part of the hydrophobic cavity for both inhibitors, although AK317 is more tightly engaged with ECL2 than AK530, explaining their similar anti-HIV-1 potencies despite the difference in Kd values. We also found that amino acid residues in the β-hairpin structural motif of ECL2 are critical for HIV-1-elicited fusion and binding of the spirodiketopiperazine-based inhibitors to CCR5. The direct ECL2-engaging property of the inhibitors likely produces an ECL2 conformation, which HIV-1 gp120 cannot bind to, but also prohibits HIV-1 from utilizing the “inhibitor-bound” CCR5 for cellular entry—a mechanism of HIV-1's resistance to CCR5 inhibitors. The data should not only help delineate the dynamics of CCR5 following inhibitor binding but also aid in designing CCR5 inhibitors that are more potent against HIV-1 and prevent or delay the emergence of resistant HIV-1 variants. 相似文献
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del Sol A Fujihashi H Amoros D Nussinov R 《Protein science : a publication of the Protein Society》2006,15(9):2120-2128
The representation of protein structures as small-world networks facilitates the search for topological determinants, which may relate to functionally important residues. Here, we aimed to investigate the performance of residue centrality, viewed as a family fold characteristic, in identifying functionally important residues in protein families. Our study is based on 46 families, including 29 enzyme and 17 non-enzyme families. A total of 80% of these central positions corresponded to active site residues or residues in direct contact with these sites. For enzyme families, this percentage increased to 91%, while for non-enzyme families the percentage decreased substantially to 48%. A total of 70% of these central positions are located in catalytic sites in the enzyme families, 64% are in hetero-atom binding sites in those families binding hetero-atoms, and only 16% belong to protein-protein interfaces in families with protein-protein interaction data. These differences reflect the active site shape: enzyme active sites locate in surface clefts, hetero-atom binding residues are in deep cavities, while protein-protein interactions involve a more planar configuration. On the other hand, not all surface cavities or clefts are comprised of central residues. Thus, closeness centrality identifies functionally important residues in enzymes. While here we focus on binding sites, we expect to identify key residues for the integration and transmission of the information to the rest of the protein, reflecting the relationship between fold and function. Residue centrality is more conserved than the protein sequence, emphasizing the robustness of protein structures. 相似文献