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Transgene integration into the human AAVS1 locus enhances myosin II-dependent contractile force by reducing expression of myosin binding subunit 85

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Title: Transgene integration into the human AAVS1 locus enhances myosin II-dependent contractile force by reducing expression of myosin binding subunit 85
Authors: Mizutani, Takeomi Browse this author →KAKEN DB
Li, Rui Browse this author
Haga, Hisashi Browse this author
Kawabata, Kazushige Browse this author
Keywords: Myosin binding subunit 85
Adeno-associated virus site 1
Cellular contractile force
Myosin regulatory light chain
phosphorylation
Genome editing
Issue Date: 18-Sep-2015
Publisher: Elsevier
Journal Title: Biochemical and biophysical research communications
Volume: 465
Issue: 2
Start Page: 270
End Page: 274
Publisher DOI: 10.1016/j.bbrc.2015.08.018
PMID: 26260320
Abstract: The adeno-associated virus site 1 (AAVS1) locus in the human genome is a strong candidate for gene therapy by insertion of an exogenous gene into the locus. The AAVS1 locus includes the coding region for myosin binding subunit 85 (MBS85). Although the function of MBS85 is not well understood, myosin II-dependent contractile force may be affected by altered expression of MBS85. The effect of altered expression of MBS85 on cellular contractile force should be examined prior to the application of gene therapy. In this study, we show that transgene integration into AAVS1 and consequent reduction of MBS85 expression changes myosin II-dependent cellular contractile force. We established a human fibroblast cell line with exogenous DNA knocked-in to AAVSI (KI cells) using the CRISPR/Cas9 genome editing system. Western blotting analysis showed that KI cells had significantly reduced MBS85 expression. KI cells also showed greater cellular contractile force than control cells. The increased contractile force was associated with phosphorylation of the myosin II regulatory light chain (MRLC). Transfection of KI cells with an MBS85 expression plasmid restored cellular contractile force and phosphorylation of MRLC to the levels in control cells. These data suggest that transgene integration into the human AAVS1 locus induces an increase in cellular contractile force and thus should be considered as a gene therapy to effect changes in cellular contractile force.
Rights: ©2015. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/
http://creativecommons.org/licenses/by-nc-nd/4.0/
Type: article (author version)
URI: http://hdl.handle.net/2115/62852
Appears in Collections:生命科学院・先端生命科学研究院 (Graduate School of Life Science / Faculty of Advanced Life Science) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)

Submitter: 水谷 武臣

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