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Quantitative analysis of condensation/decondensation status of pDNA in the nuclear sub-domains by QD-FRET

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Title: Quantitative analysis of condensation/decondensation status of pDNA in the nuclear sub-domains by QD-FRET
Authors: Shaheen, Sharif M. Browse this author
Akita, Hidetaka Browse this author
Yamashita, Atsushi Browse this author
Katoono, Ryo Browse this author
Yui, Nobuhiko Browse this author
Biju, Vasudevanpillai Browse this author
Ishikawa, Mitsuru Browse this author
Harashima, Hideyoshi Browse this author
Issue Date: Apr-2011
Publisher: Oxford University Press
Journal Title: Nucleic Acids Research
Volume: 39
Issue: 7
Start Page: e48
Publisher DOI: 10.1093/nar/gkq1327
Abstract: Recent studies indicate that controlling the nuclear decondensation and intra-nuclear localization of plasmid DNA (pDNA) would result in an increased transfection efficiency. In the present study, we established a technology for imaging the nuclear condensation/decondensation status of pDNA in nuclear subdomains using fluorescence resonance energy transfer (FRET) between quantum dot (QD)-labeled pDNA as donor, and rhodamine-labeled polycations as acceptor. The FRET-occurring pDNA/polycation particle was encapsulated in a nuclear delivery system; a tetra-lamellar multifunctional envelope-type nano device (T-MEND), designed to overcome the endosomal membrane and nuclear membrane via step-wise fusion. Nuclear subdomains (i.e. heterochromatin and euchromatin) were distinguished by Hoechst33342 staining. Thereafter, Z-series of confocal images were captured by confocal laser scanning microscopy. pDNA in condensation/decondensation status in heterochromatin or euchromatin were quantified based on the pixel area of the signals derived from the QD and rhodamine. The results obtained indicate that modulation of the supra-molecular structure of polyrotaxane (DMAE-ss-PRX), a condenser that is cleaved in a reductive environment, conferred euchromatin-preferred decondensation. This represents the first demonstration of the successful control of condensation/decondensation in specific nuclear sub-domain via the use of an artificial DNA condenser.
Type: article
Appears in Collections:薬学研究院 (Faculty of Pharmaceutical Sciences) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)

Submitter: 原島 秀吉

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