Complete Nucleotide Sequence and Genetic Organization of Aichi Virus, a Distinct Member of the Picornaviridae Associated with Acute Gastroenteritis in Humans

The complete nucleotide sequence of a novel enteric virus, Aichi virus, associated with nonbacterial acute gastroenteritis in humans was determined. The Aichi virus genome proved to be a single-stranded positive-sense RNA molecule with 8,251 bases excluding a poly(A) tail; it contains a large open reading frame with 7,302 nucleotides that encodes a potential polyprotein precursor of 2,433 amino acids. The genome contains a 5′ nontranslated region (NTR) with 712 bases and a 3′ NTR with 240 bases followed by a poly(A) tail. The structure of the genome, VPg–5′ NTR–leader protein–structural proteins–nonstructural proteins–3′ NTR–poly(A), was found to be typical of a picornavirus. The VP0-VP3 and VP3-VP1 cleavage sites were determined to be Q-H and Q-T, respectively, by N-terminal amino acid sequence analyses using purified virion proteins. Possible cleavage sites, Q-G, Q-A, and Q-S, which cleave P2 and P3 polyproteins were found to be similar to those of picornaviruses. A dendrogram based on 3D^pol proteins indicated that Aichi virus is genetically distinct from the known six genera of picornaviruses including entero-, rhino-, cardio-, aphtho-, and hepatovirus and echovirus 22. Considering this together with other properties of the virus (T. Yamashita, S. Kobayashi, K. Sakae, S. Nakata, S. Chiba, Y. Ishihara, and S. Isomura, J. Infect. Dis. 164:954–957, 1991), we propose that Aichi virus be regarded as a new genus of the family Picornaviridae.     

Multiplication and Differentiation of Plastid Nucleoids during Development of Chloroplasts and Etioplasts from Proplastids in Triticum aestivum

The morphological changes of plastid nucleoids (pt nucleoids)in the shoot apex and along the axis of the leaf blade in Triticumaestivum L. cv. Asakaze were followed with fluorescence microscopyafter staining with 4'6-diamidino-2-phenylindole (DAPI) andquantified with supersensitive microspectrophotometry. Proplastidsin the shoot apex contained 1–10 spherical pt nucleoids.These pt nucleoids changed to a row of spherical and cup-shapedpt nucleoids in sausage-shaped plastids at the leaf base inboth dark and light conditions, in which active cell divisionwas observed. These structures have a higher copy number ofplastid DNA (pt DNA) (72–78 copies) compared to proplastidsin the shoot apex (32–45 copies) and, therefore, may reflectthat active pt DNA synthesis is in progression. In the dark,the cup-shaped pt nucleoids in the spherical etioplasts, whichoriginated from the sausage-shaped plastids, grew to form ring-shapedpt nucleoids. Each ring-shaped pt nucleoid is sub-divided intosmaller pt nucleoids. Under continuous illumination, similarmorphological changes of pt nucleoids occurred except for distributionof small pt nucleoids into young chloroplasts as well as inmature chloroplasts. However, pt nucleotids of leucoplasts inepidermal and vascular bundle sheath cells did not show conspicuouschanges along the axis of the leaf blade. The significance ofthese observations is discussed in relation to plastid differentiationand to the plastid division cycle. ⁴ Present address: Faculty of Science, University of Tokyo,Hongo, Bunkyo-ku, Tokyo, 113 (Received August 15, 1989; Accepted April 13, 1990)     

Design,synthesis and evaluation of γ-turn mimetics as LSD1-selective inhibitors

Lysine-specific demethylase 1 (LSD1) is an attractive molecular target for cancer therapy. We have previously reported potent LSD1-selective inhibitors (i.e., NCD18, NCD38, and their analogs) consisting of trans-2-phenylcyclopropylamine (PCPA) or trans-2-arylcyclopropylamine (ACPA) and a lysine moiety that could form a γ-turn structure in the active site of LSD1. Herein we report the design, synthesis and evaluation of γ-turn mimetic compounds for further improvement of LSD1 inhibitory activity and anticancer activity. Among a series of γ-turn mimetic compounds synthesized by a Mitsunobu-reaction-based amination strategy, we identified 1n as a potent and selective LSD1 inhibitor. Compound 1n induced cell cycle arrest and apoptosis through histone methylation in human lung cancer cells. The γ-turn mimetics approach should offer new insights into drug design for LSD1-selective inhibitors.     

Expression and self-assembly of empty virus-like particles of hepatitis E virus.

Hepatitis E virus (HEV) is a pathogenic agent that causes fecally-orally transmitted acute hepatitis. The genome, a single-stranded positive-sense RNA, encodes three forward open reading frames (ORFs), in which an approximately 2-kb structural protein is located in the 3' end. To produce HEV-like particles the structural protein, with its N terminus truncated (amino acid residues 112 to 660 of ORF2), was expressed in insect Tn5 cells by a recombinant baculovirus. In addition to the primary translation product with a molecular mass of 58 kDa, a large amount of a further-processed molecule with a molecular mass of 50 kDa was generated and efficiently released into the culture medium. Electron microscopic observation of the culture medium revealed that the 50-kDa protein self-assembled to form empty virus-like particles (VLPs). The buoyant density of the VLPs in CsCl was 1.285 g/cm3 and their diameter was 23.7 nm, a little smaller than the 27 nm of native HEV particles secreted into the bile or stools of experimentally infected monkeys. The yield of the VLPs was 1 mg per 10(7) cells as a purified form. The particles possess antigenicity similar to that of authentic HEV particles and, consequently, they appear to be a good antigen for the sensitive detection of HEV-specific immunoglobulin G (IgG) and IgM antibodies. Furthermore, the VLP may be the most promising candidate yet for an HEV vaccine, owing to its potent immunogenicity.     

Disappearance of plastid and mitochondrial nucleoids during the formation of generative cells of higher plants revealed by fluorescence microscopy

Summary The fate of plastid and mitochondrial nucleoids (pt and mt nucleoids) ofTriticum aestivum was followed during the reproductive organ formation using fluorescence microscopy after staining with 4'6-diamidino-2-phenylindole (DAPI). This investigation showed a drastic morphological change of pt nucleoids during the differentiation of reproductive organs from the shoot apex. Dot-shaped pt nucleoids grew into ring-shaped ones, which divided into small pieces in the monocellular pollen grain, as observed in this plant's earlier stage of leaf development. During the development of mature pollen grain from monocellular pollen grain, pt and/or mt nucleoids disappeared through the division of the male generative cell ofT. aestivum. Cytologically, this observation is direct evidence of the maternal inheritance of higher plants. Thus far, cytological evidence of this phenomenon has been found mostly by morphological criteria using electron microscopy, which admits some ambiguity. In the plants exemplified byLilium longiflorum, pt and/or mt nucleoids disappeared after the first pollen grain mitosis, which precededT. aestivum. In the plants exemplified byTrifolium repens, pt and/or mt nucleoids existed in the generative cells of the mature pollen grain.The significance of these observations was discussed in relation to the interaction between nuclear and organelle genomes during plant development.Abbreviations DAPI 4'6 diamidino-2-phenylindole - Mt DNA Mitochondrial DNA - Mt nucleoid Mitochondrial nucleoid - Pt DNA Plastid DNA - Pt nucleoid Plastid nucleoid On leave from Department of Biology, Nagoya University, Furocho, Chikusaku, Nagoya 464, Japan.     

Cross transfer effects of muscular training on blood flow in the ipsilateral and contralateral forearms

Blood flow in the right and left forearms was determined by venous occlusion plethysmography in ten healthy male subjects before and after training with a hand ergometer. The subjects in group A and B were trained using work loads of 1/3 and 1/2, respectively, of maximum grip strength 6 days/week for 6 weeks. It was found that the blood flow in the left (untrained or contralateral) forearm during exhaustive training of the right hand increased gradually with increasing training periods, and that after 6 weeks of training, grip strength, endurance and peak blood flow of the forearm increased significantly not only in the trained forearm, but also in the untrained forearm. From these results, it is suggested that the increase of blood flow in the contralateral limb after training may, at least in part, be related to the cross transfer effect of muscular endurance.     

Phenotype of polyoma-induced hamster tumor cells lines.

Cell lines from polyoma-induced hamster tumors exhibit a fully transformed phenotype despite the absence of the 105K (105,000-dalton) form of polyoma T-antigen.     

Characterization of JC papovavirus adapted to growth in human embryonic kidney cells.

Human papovavirus JC virus was adapted to growth in human embryonic kidney (HEK) cells. After eight passages, the HEK-adapted JC virus produced high virus yields and was capable of forming plaques in HEK monolayer cultures. Eleven plaque-purified stocks were prepared and characterized. Biologically, the plaque-purified virus induced tumor and viral antigens in HEK cells earlier and in a higher percentage of cells than uncloned virus. Cytopathic changes were also evident sooner and were more extensive. The DNA from uncloned as well as plaque-purified isolates was analyzed by restriction endonuclease cleavage followed by gel electrophoresis. The DNA from uncloned HEK-adapted virus was heterogeneous. Plaque-purified virus isolates yielded DNA which, although much less heterogeneous than the uncloned stock, still consisted of two or more species of viral DNA.