Single Quantum Dot Tracking Reveals that an Individual Multivalent HIV-1 Tat Protein Transduction Domain Can Activate Machinery for Lateral Transport and Endocytosis

Yasuhiro Suzuki, Chandra Nath Roy, Warunya Promjunyakul, Hiroyasu Hatakeyama, Kohsuke Gonda, Junji Imamura, Biju Vasudevanpillai, Noriaki Ohuchi, Makoto Kanzaki, Hideo Higuchi, Mitsuo Kaku

Research output: Contribution to journalArticlepeer-review

17 Citations (Scopus)

Abstract

The mechanisms underlying the cellular entry of the HIV-1 Tat protein transduction domain (TatP) and the molecular information necessary to improve the transduction efficiency of TatP remain unclear due to the technical limitations for direct visualization of TatP's behavior in cells. Using confocal microscopy, total internal reflection fluorescence microscopy, and four-dimensional microscopy, we developed a single-molecule tracking assay for TatP labeled with quantum dots (QDs) to examine the kinetics of TatP initially and immediately before, at the beginning of, and immediately after entry into living cells. We report that even when the number of multivalent TatP (mTatP)-QDs bound to a cell was low, each single mTatP-QD first locally induced the cell's lateral transport machinery to move the mTatP-QD toward the center of the cell body upon cross-linking of heparan sulfate proteoglycans. The centripetal and lateral movements were linked to the integrity and flow of actomyosin and microtubules. Individual mTatP underwent lipid raft-mediated temporal confinement, followed by complete immobilization, which ultimately led to endocytotic internalization. However, bivalent TatP did not sufficiently promote either cell surface movement or internalization. Together, these findings provide clues regarding the mechanisms of TatP cell entry and indicate that increasing the valence of TatP on nanoparticles allows them to behave as cargo delivery nanomachines.

Original languageEnglish
Pages (from-to)3036-3049
Number of pages14
JournalMolecular and cellular biology
Volume33
Issue number15
DOIs
Publication statusPublished - 2013 Aug

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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