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16 Cell-Penetrating Peptides: Processes and Applications
1.4.4 CELLULAR ASPECTS OF TAT CPP UPTAKE
The internalization of Tat-derived CPP is not cell specific because all cell lines tested
so far in our laboratory efficiently entrapped the native Tat-translocating peptide.
Additional data in the literature confirmed the broad panel of cells allowing Tat
peptide uptake. These include cell types such as monocyte or macrophage progenitors
that are poorly transfected by traditional methods. 53 Importantly, Tat peptideconjugated
molecules were also shown to pass through the blood–brain barrier,
which is a highly selective biological membrane; 36 Tat-derived peptides, such as a
short homopolymer of arginine, efficiently cross the skin stratum corneum. 54
Tat uptake by these different cell types most probably involves a common cellular
component or a common process of entry that still remains to be highlighted.
However, some indications from the literature raise interesting features about Tat
(and CPP-related peptides) cellular uptake. The absence of perinuclear punctuation
after incubation of cells with fluorescence-labeled Tat peptides (References 23, for
native Tat CPP, and 47 for Tat-derived peptide) and the important uptake at low
temperature excluded the endocytosis pathway, as previously discussed. Pinocytosis
and potocytosis, a caveolae-mediated uptake (see References 55 and 56 for recent
reviews), could be involved in the translocation of the Tat peptide and its derivatives,
since potocytosis is poorly sensitive to temperature. 57 In line with this possibility, a
reduction of the uptake of a Tat peptide-displaying phage was observed in the
presence of inhibitors of caveolae formation such as nystatin or fillipin. 39 However,
other inhibitors of caveolae formation such as okadaic acid did not inhibit the uptake
of a fluorescein-labeled Tat CPP. 23 The huge difference in the nature and size of
these Tat-conjugated entities (a bacteriophage vs. a small peptide) could explain
these apparent contradictory results; the possible involvement of caveolae in CPP
uptake remains to be convincingly established.
It was shown that this Tat CPP peptide was able to vectorize various cargo
molecules (from small peptide to phage or ferromagnetic particles) inside
cells. 14,15,27,28,33,36,39,58-62 Whether a single pathway is involved in all cases has yet to
be evaluated. Whether binding to other cell surface determinants (for instance, to
polar lipid heads) is required has also to be further investigated. Along this line, the
requirement of an appropriate exposure of the Tat peptide at the surface of liposomes
to allow their cellular uptake has been highlighted. 42 This feature strongly suggests
that a direct interaction with unknown cellular entities is requested to trigger the
internalization process.
1.5 CONCLUSION AND PERSPECTIVES
As discussed in Section 1.4 and other chapters in this book, short natural or synthetic
basic peptides unexpectedly allowed a mechanism not involving cell surface receptors
and not mediated by endocytosis to cross biological barriers. Whether the
formation of inverted micelles, as proposed for the Antennapedia peptide, 63 or
caveolae might be involved as suggested by Eguchi et al. 39 is the object of ongoing
experiments.