title: Retargeting of microcell fusion towards recipient cell-oriented transfer of human artificial chromosome.
abstract: BACKGROUND
  Human artificial chromosome (HAC) vectors have some unique characteristics as compared with conventional vectors, carrying large transgenes without size limitation, showing persistent expression of transgenes, and existing independently from host genome in cells. With these features, HACs are expected to be promising vectors for modifications of a variety of cell types. However, the method of introduction of HACs into target cells is confined to microcell-mediated chromosome transfer (MMCT), which is less efficient than other methods of vector introduction. Application of Measles Virus (MV) fusogenic proteins to MMCT instead of polyethylene glycol (PEG) has partly solved this drawback, whereas the tropism of MV fusogenic proteins is restricted to human CD46- or SLAM-positive cells.
  
  
  RESULTS
  Here, we show that retargeting of microcell fusion by adding anti-Transferrin receptor (TfR) single chain antibodies (scFvs) to the extracellular C-terminus of the MV-H protein improves the efficiency of MV-MMCT to human fibroblasts which originally barely express both native MV receptors, and are therefore resistant to MV-MMCT. Efficacy of chimeric fusogenic proteins was evaluated by the evidence that the HAC, tagged with a drug-resistant gene and an EGFP gene, was transferred from CHO donor cells into human fibroblasts. Furthermore, it was demonstrated that no perturbation of either the HAC status or the functions of transgenes was observed on account of retargeted MV-MMCT when another HAC carrying four reprogramming factors (iHAC) was transferred into human fibroblasts.
  
  
  CONCLUSIONS
  Retargeted MV-MMCT using chimeric H protein with scFvs succeeded in extending the cell spectrum for gene transfer via HAC vectors. Therefore, this technology could facilitate the systematic cell engineering by HACs.
authors.affiliation: Division of Molecular and Cell Genetics, Department of Molecular and Cellular Biology, School of Life Sciences, Faculty of Medicine, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. mhiratsu@med.tottori-u.ac.jp.
authors.forename: Masaharu
authors.initials: M
authors.lastname: Hiratsuka
authors.affiliation: Division of Molecular Genetics and Biofunction, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. ueda@chromocenter.com.
authors.forename: Kana
authors.initials: K
authors.lastname: Ueda
authors.affiliation: Division of Molecular Genetics and Biofunction, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. narumi@med.tottori-u.ac.jp.|Chromosome Engineering Research Center, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. narumi@med.tottori-u.ac.jp.
authors.forename: Narumi
authors.initials: N
authors.lastname: Uno
authors.affiliation: Division of Molecular Genetics and Biofunction, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. katsuhiro112@med.tottori-u.ac.jp.
authors.forename: Katsuhiro
authors.initials: K
authors.lastname: Uno
authors.affiliation: Division of Molecular Genetics and Biofunction, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. syk3a_419@yahoo.co.jp.
authors.forename: Sayaka
authors.initials: S
authors.lastname: Fukuhara
authors.affiliation: Division of Integrative Bioscience, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, Japan. hkurosaki@grape.med.tottori-u.ac.jp.
authors.forename: Hajime
authors.initials: H
authors.lastname: Kurosaki
authors.affiliation: Division of Molecular Genetics and Biofunction, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. shoko0206@grape.med.tottori-u.ac.jp.
authors.forename: Shoko
authors.initials: S
authors.lastname: Takehara
authors.affiliation: Division of Pathological Biochemistry, Department of Biomedical Sciences, School of Life Sciences, Faculty of Medicine, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. osamitsu@grape.med.tottori-u.ac.jp.
authors.forename: Mitsuhiko
authors.initials: M
authors.lastname: Osaki
authors.affiliation: Division of Molecular Genetics and Biofunction, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. kazuki@grape.med.tottori-u.ac.jp.|Chromosome Engineering Research Center, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. kazuki@grape.med.tottori-u.ac.jp.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kazuki
authors.affiliation: Division of Antibody Project, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi, 470-1192, Japan. kurosawa@fujita-hu.ac.jp.
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Kurosawa
authors.affiliation: Division of Integrative Bioscience, Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, Japan. taka@med.tottori-u.ac.jp.
authors.forename: Takafumi
authors.initials: T
authors.lastname: Nakamura
authors.affiliation: Division of Human Genome Science, Department of Molecular and Cellular Biology, School of Life Sciences, Faculty of Medicine, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. mkatoh@grape.med.tottori-u.ac.jp.
authors.forename: Motonobu
authors.initials: M
authors.lastname: Katoh
authors.affiliation: Chromosome Engineering Research Center, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan. oshimura@med.tottori-u.ac.jp.|Japan Science and Technology Agency, CREST, 5, Sanbancho, Chiyoda-ku, Tokyo, 102-0075, Japan. oshimura@med.tottori-u.ac.jp.
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Oshimura
mesh_terms: D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D022222:Chromosomes, Artificial, Human / Q000235:genetics*; D006224:Cricetinae; D003412:Cricetulus; D005347:Fibroblasts; D018014:Gene Transfer Techniques; D006801:Humans; D008459:Measles virus / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014760:Viral Fusion Proteins / Q000235:genetics*
chemical_list: D011993:Recombinant Fusion Proteins; D014760:Viral Fusion Proteins