In vivo protein transduction: intracellular delivery of biologically active proteins, compounds and DNA

Schwarze, Steven R.; Dowdy, Steven F.

doi:10.1016/s0165-6147(99)01429-7

Cited by 422 publications

(297 citation statements)

References 24 publications

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“…In contrast to this, the chosen TAT-transduction system overcomes the limitations that could occur in case of DNA transfection such as low transfection efficiency. The TATtransduction technique allows a fast and concentration-dependent delivery of TAT fusion proteins into nearly 100% of cells, where they remain for 48 h until their degradation (Schwarze and Dowdy, 2000;Abu-Amer et al, 2001). Using the TAT protein transduction technology, protein purification can be performed regardless of renaturation or folding since misfolding enhances the uptake of the protein into the cells where it gets folded correctly (Nagahara et al, 1998).…”

Section: Discussionmentioning

confidence: 99%

Antitumour effects of PLC-γ1-(SH2)2-TAT fusion proteins on EGFR/c-erbB-2-positive breast cancer cells

et al. 2004

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Due to its pivotal role in the growth factor-mediated tumour cell migration, the adaptor protein phospholipase C-g1 (PLC-g1) is an appropriate target to block ultimately the spreading of EGFR/c-erbB-2-positive tumour cells, thereby minimising metastasis formation. Here, we present an approach to block PLC-g1 activity by using protein-based PLC-g1 inhibitors consisting of PLC-g1 SH2 domains, which were fused to the TAT-transduction domain to ensure a high protein transduction efficiency. Two proteins were generated containing one PLC-g1-SH2-domain (PS1-TAT) or two PLC-g1-SH2 domains (PS2-TAT). PS2-TAT treatment of the EGFR/c-erbB-2-positive cell line MDA-HER2 resulted in a reduction of the EGF-mediated PLC-g1 tyrosine phosphorylation of about 30%, concomitant with a complete abrogation of the EGF-driven calcium influx. In addition to this, long-term PS2-TAT treatment both reduces the EGF-mediated migration of about 75% combined with a markedly decreased time locomotion of single MDA-HER2 cells as well as decreases the proliferation of MDA-HER2 cells by about 50%. Due to its antitumoral capacity on EGFR/c-erbB-2-positive breast cancer cells, we conclude from our results that the protein-based PLC-g1 inhibitor PS2-TAT may be a means for novel adjuvant antitumour strategies to minimise metastasis formation because of the blockade of cell migration and proliferation.

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Section: Discussionmentioning

confidence: 99%

Antitumour effects of PLC-γ1-(SH2)2-TAT fusion proteins on EGFR/c-erbB-2-positive breast cancer cells

et al. 2004

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“…Recently, methods have been developed for the delivery of exogenous proteins into living cells with the help of membranepermeable carrier peptides such as HIV-1 1 Tat-(48 -60) and Antennapedia-(43-58) (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11). By genetically or chemically hybridizing these carrier peptides, the efficient intracellular delivery of various oligopeptides and proteins was achieved.…”

mentioning

confidence: 99%

Arginine-rich Peptides

Futaki

Suzuki

Ohashi

et al. 2001

Journal of Biological Chemistry

1,498

591

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A basic peptide derived from human immunodeficiency virus (HIV)-1 Tat protein (positions 48 -60) has been reported to have the ability to translocate through the cell membranes and accumulate in the nucleus, the characteristics of which are utilized for the delivery of exogenous proteins into cells. Based on the fluorescence microscopic observations of mouse macrophage RAW264.7 cells, we found that various arginine-rich peptides have a translocation activity very similar to Tat-(48 -60). These included such peptides as the D-amino acid-and arginine-substituted Tat-(48 -60), the RNA-binding peptides derived from virus proteins, such as HIV-1 Rev, and flock house virus coat proteins, and the DNA binding segments of leucine zipper proteins, such as cancer-related proteins c-Fos and c-Jun, and the yeast transcription factor GCN4. These segments have no specific primary and secondary structures in common except that they have several arginine residues in the sequences. Moreover, these peptides were able to be internalized even at 4°C. These results strongly suggested the possible existence of a common internalization mechanism ubiquitous to arginine-rich peptides, which is not explained by a typical endocytosis. Using (Arg) n (n ‫؍‬ 4 -16) peptides, we also demonstrated that there would be an optimal number of arginine residues (n ϳ 8) for the efficient translocation.Recently, methods have been developed for the delivery of exogenous proteins into living cells with the help of membranepermeable carrier peptides such as HIV-1 1 Tat-(48 -60) and Antennapedia-(43-58) (1-11). By genetically or chemically hybridizing these carrier peptides, the efficient intracellular delivery of various oligopeptides and proteins was achieved. One of the most amazing examples is the Tat-␤-galactosidase fusion protein (4), which has a molecular mass as high as 120 kDa.Intraperitoneal injection of the protein resulted in delivery of the protein with ␤-galactosidase activity to various tissues in mice, including the brain. The peptide-mediated approaches would allow the incorporation of peptides containing unnatural amino acids or nonpeptide molecules such as fluorescence probes. These methods would become powerful tools not only for therapeutic purposes as an alternative to gene delivery, but also for the understanding of the mechanisms behind fundamental cellular events, such as signal transduction and gene transcription.Besides the potential of Tat-(48 -60) as a protein carrier, the internalization mechanism of the peptide attracted our interest. For example, Tat-(48 -60) (GRKKRRQRRRPPQ) is a highly basic and hydrophilic peptide, which contains 6 arginine and 2 lysine residues in its 13 amino acid residues. However, the peptide was reported to be translocated through the cell membranes in 5 min at a concentration of 0.1 M (2). Internalization of the peptide was not inhibited even at 4°C. The peptide is less toxic to cells than other basic membrane-interacting agents. The above features suggested that the internalization mechanism of Tat-...

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“…It can deliver a wide variety of proteins across the plasma membrane by a mechanism referred to as "protein transduction" (8,9). An inherent characteristic of the specific sequences or numbers of the amino acids arginine or lysine, or some structural motif, may be important in process of protein transduction (10,11). Thus, it might be a useful molecular tool for achieving the widespread distribution of recombinant proteins in gene therapy applications (10).…”

Section: Enhancement Of Adenoviral Transduction and Immunogenecity Ofmentioning

confidence: 99%

“…An inherent characteristic of the specific sequences or numbers of the amino acids arginine or lysine, or some structural motif, may be important in process of protein transduction (10,11). Thus, it might be a useful molecular tool for achieving the widespread distribution of recombinant proteins in gene therapy applications (10). Several groups have previously demonstrated the endogenous expression of a TAT fusion protein in vitro, followed by intercellular trafficking (12).…”

Section: Enhancement Of Adenoviral Transduction and Immunogenecity Ofmentioning

confidence: 99%

Enhancement of Adenoviral Transduction and Immunogenecity of Transgenes by Soluble Coxsackie and Adenovirus Receptor-TAT Fusion Protein on Dendritic Cells

et al. 2006

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Background: Investigating strategy to enhance efficiency of gene transfer via adenovirus is critical to sustain gene expression in targeted cells or tissues to regulate immune responses. However, the use of adenovirus as a gene delivery method has been limited by the native tropism of the virus. In this study, the critical parameter is to improve the efficient binding of viral particles to the plasma membrane prior to cellular uptake.

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In vivo protein transduction: intracellular delivery of biologically active proteins, compounds and DNA

Cited by 422 publications

References 24 publications

Antitumour effects of PLC-γ1-(SH2)2-TAT fusion proteins on EGFR/c-erbB-2-positive breast cancer cells

Antitumour effects of PLC-γ1-(SH2)2-TAT fusion proteins on EGFR/c-erbB-2-positive breast cancer cells

Arginine-rich Peptides

Enhancement of Adenoviral Transduction and Immunogenecity of Transgenes by Soluble Coxsackie and Adenovirus Receptor-TAT Fusion Protein on Dendritic Cells

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