Organelle-Targeted Delivery of Biological Macromolecules Using the Protein Transduction Domain: Potential Applications for Peptide Aptamer Delivery into the Nucleus

• Published in: Journal of molecular biology Vol. 380; no. 5; pp. 777 - 782
• Main Authors: Yoshikawa, Tomoaki, Sugita, Toshiki, Mukai, Yohei, Yamanada, Natsue, Nagano, Kazuya, Nabeshi, Hiromi, Yoshioka, Yasuo, Nakagawa, Shinsaku, Abe, Yasuhiro, Kamada, Haruhiko, Tsunoda, Shin-ichi, Tsutsumi, Yasuo
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 25.07.2008
• Subjects: Aptamers, Peptide - genetics; Aptamers, Peptide - metabolism; Base Sequence; Cell Line; Cell Nucleus - chemistry; Cell Nucleus - metabolism; Cell Survival - drug effects; Drug Delivery Systems - methods; Drug Design; Gene Products, tat - chemistry; HA2; HeLa Cells; HIV-1 - chemistry; Humans; intracellular targeting; Molecular Sequence Data; nuclear localization signal; Nuclear Localization Signals; peptide aptamer; Peptides - chemistry; Peptides - metabolism; Peptides - pharmacology; Protein Structure, Tertiary; Protein Transport - drug effects; Protein Transport - physiology; Recombinant Fusion Proteins - metabolism; Sequence Analysis, DNA; Tat; Time Factors; PTD; PM10–Tat; Tat; Tat–cargo; nuclear localization signal; HA2–Tat; NLS–PM10–Tat; peptide aptamer; NLS; MDM2; HA2; intracellular targeting; NLS–VENUS–Tat
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2008.05.047

Extensive effort is currently being expended on the innovative design and engineering of new molecular carrier systems for the organelle-targeted delivery of biological cargoes (e.g., peptide aptamers or biological proteins) as tools in cell biology and for developing novel therapeutic approaches. Although cell-permeable Tat peptides are useful carriers for delivering biological molecules into the cell, much internalized Tat-fused cargo is trapped within macropinosomes and thus not delivered into organelles. Here, we devised a novel intracellular targeting technique to deliver Tat-fused cargo into the nucleus using an endosome-disruptive peptide (hemagglutinin-2 subunit) and a nuclear localization signal peptide. We show for the first time that Tat-conjugated peptide aptamers can be selectively delivered to the nucleus by using combined hemagglutinin-2 subunit and nuclear localization signal peptides. This nuclear targeting technique resulted in marked enhancement of the cytostatic activity of a Tat-fused p53-derived peptide aptamer against human MDM2 (mouse double minute 2) that inhibits p53–MDM2 binding. Thus, our technique provides a unique methodology for the development of novel therapeutic approaches based on intracellular targeting.