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      Identification of the NLS and NES motifs of VP2 from chicken anemia virus and the interaction of VP2 with mini-chromosome maintenance protein 3

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          Abstract

          Background

          VP2 of chicken anemia virus (CAV) is a dual-specificity phosphatase required for virus infection, assembly and replication. The functions of the nuclear localization signal (NLS) and nuclear export signal (NES) of VP2 in the cell, however, are poorly understood. Our study identified the presence of a NLS in VP2 and showed that the protein interacted significantly with mini-chromosome maintenance protein 3 (MCM3) in the cell.

          Results

          An arginine-lysine rich NLS could be predicted by software and spanned from amino acids 133 to 138 of VP2. The critical amino acids residues between positions 136 and 138, and either residue 133 or 134 are important for nuclear import in mammalian cells based on systematic mutagenesis. A NES is also predicted in VP2; however the results suggest that no functional NES is present and that this protein is CRM1 independent. It was also shown that VP2 is a chromatin binding protein and, notably, using a co-immunoprecipitation assay, it was found that VP2 association with MCM3 and that this interaction does not require DSP activity.

          Conclusions

          VP2 contains a NLS that span from amino acids 133 to 138. VP2 is a CRM1 independent protein during nuclear export and associates with MCM3 in cells.

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          Most cited references35

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          NLStradamus: a simple Hidden Markov Model for nuclear localization signal prediction

          Background Nuclear localization signals (NLSs) are stretches of residues within a protein that are important for the regulated nuclear import of the protein. Of the many import pathways that exist in yeast, the best characterized is termed the 'classical' NLS pathway. The classical NLS contains specific patterns of basic residues and computational methods have been designed to predict the location of these motifs on proteins. The consensus sequences, or patterns, for the other import pathways are less well-understood. Results In this paper, we present an analysis of characterized NLSs in yeast, and find, despite the large number of nuclear import pathways, that NLSs seem to show similar patterns of amino acid residues. We test current prediction methods and observe a low true positive rate. We therefore suggest an approach using hidden Markov models (HMMs) to predict novel NLSs in proteins. We show that our method is able to consistently find 37% of the NLSs with a low false positive rate and that our method retains its true positive rate outside of the yeast data set used for the training parameters. Conclusion Our implementation of this model, NLStradamus, is made available at:
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            MCM2-7 complexes bind chromatin in a distributed pattern surrounding the origin recognition complex in Xenopus egg extracts.

            The MCM2-7 complex is believed to function as the eukaryotic replicative DNA helicase. It is recruited to chromatin by the origin recognition complex (ORC), Cdc6, and Cdt1, and it is activated at the G(1)/S transition by Cdc45 and the protein kinases Cdc7 and Cdk2. Paradoxically, the number of chromatin-bound MCM complexes greatly exceeds the number of bound ORC complexes. To understand how the high MCM2-7:ORC ratio comes about, we examined the binding of these proteins to immobilized linear DNA fragments in Xenopus egg extracts. The minimum length of DNA required to recruit ORC and MCM2-7 was approximately 80 bp, and the MCM2-7:ORC ratio on this fragment was approximately 1:1. With longer DNA fragments, the MCM2-7:ORC ratio increased dramatically, indicating that MCM complexes normally become distributed over a large region of DNA surrounding ORC. Only a small subset of the chromatin-bound MCM2-7 complexes recruited Cdc45 at the onset of DNA replication, and unlike Cdc45, MCM2-7 was not limiting for DNA replication. However, all the chromatin-bound MCM complexes may be functional, because they were phosphorylated in a Cdc7-dependent fashion, and because they could be induced to support Cdk2-dependent Cdc45 loading. The data suggest that in Xenopus egg extracts, origins of replication contain multiple, distributed, initiation-competent MCM2-7 complexes.
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              Isolation and Some Characteristics of an Agent Inducing Anemia in Chicks

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                Author and article information

                Journal
                BMC Vet Res
                BMC Veterinary Research
                BioMed Central
                1746-6148
                2012
                7 February 2012
                : 8
                : 15
                Affiliations
                [1 ]Department of Medical Research, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung University College of Medicine, Kaohsiung, Taiwan
                [2 ]Department of Food Science, National Pingtung University of Science and Technology, Pingtung, Taiwan
                [3 ]Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, Taiwan
                [4 ]Department of Medical Research, Tung's Taichung Metro Harbor Hospital, Taichung, Taiwan
                [5 ]Department of Safety, Health and Environmental Engineering, Mingchi University of Technology, Taipei, Taiwan
                [6 ]School of Chinese Pharmaceutical Science and Chinese Medicine Resources, China Medical University, Taichung, Taiwan
                Article
                1746-6148-8-15
                10.1186/1746-6148-8-15
                3295642
                22309683
                76170daa-8028-4516-905d-6113cb64c098
                Copyright ©2012 Cheng et al; BioMed Central Ltd.

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 21 August 2011
                : 7 February 2012
                Categories
                Research Article

                Veterinary medicine
                Veterinary medicine

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