Uptake Mechanism of Cell-Penetrating Peptides

Abstract

Cell-penetrating peptides have been extensively used since their discovery for delivering cargoes unable to cross the cell membrane. Among other transported cargoes, they have shown very efficient delivery for oligonucleotides making cell-penetrating peptides a powerful tool for gene therapy. Numerous cell-penetrating peptides have now been discovered offering a wide library of structures and mechanisms of actions. Nevertheless, if it is known that different pathways are available for particles to be taken up, most mechanisms by which these particles enter cells are still to be characterized more precisely. Indeed it is admitted that cell-penetrating peptides are taken up either by direct translocation or by endocytosis but classes of cell-penetrating peptides are usually not related to specific entrance mechanisms. Actually, for most particles, different pathways can be detected during their uptake which makes the literature sometimes contradictory. Recent studies have nevertheless shown convergent uptake patterns for individual structures. Acetylated cell-penetrating peptides complexed with oligonucleotides have been shown to interact to scavenger receptor class A to induce caveolae-mediated endocytosis whereas antimicrobial peptides create pores in the cell membrane for direct translocation. Arginine-rich peptides have presented concentration-dependent mechanisms, being taken up either by membrane destabilization or clathrin-mediated endocytosis. Relating the structure of cell-penetrating peptides or their particles to distinct mechanisms would allow this delivery platform to become even more specific by using rational design to fit to the desired uptake pathway.

Design and biological characterization of novel cell-penetrating peptides preferentially targeting cell nuclei and subnuclear regions

“…6 ). Hereby, energy-dependent pathways are usually suppressed and direct peptide translocation can be observed [ 36 ]. After treating MCF-7 and HeLa cells for 30 min with the peptides at 4 °C, both fusion peptides were distributed within the cytoplasm and also accumulated in the cell nuclei and nucleoli.…”

Gene delivery by peptide-assisted transport

“…Peptide-mediated transport mechanisms across the plasma membrane include uptake through endocytosis or macropinocytosis (right) [21]; active uptake through receptor-mediated endocytosis (bottom) [22]; and pore formation (left) [23]. Some peptides can mediate transcytosis through sequential endocytic uptake, subcellular transport, and exocytosis [24]. Within cells, peptides can facilitate endosomal escape [22] or nuclear targeting [25], leading to increased RNA delivery or gene transfection.…”

“…(b) Endocytic pathways of CPPs through clathrin and caveolin mediated endocytosis, and macropinocytosis (Reproduced from reference [24], with permission from Springer Nature).…”

Intracellular Delivery of DNA and Protein by a Novel Cell-Permeable Peptide Derived from DOT1L

“…This suggests that lipid raft-mediated endocytosis may not mediate intracellular delivery of Dot1l, although receptor-mediated endocytosis may be slightly involved. Therefore, we found that endocytosis may not be the major pathway involving the translocation of Dot1l peptide, even though the endocytosis pathway can mediate CPP uptake [30].…”

“…As shown in Figure 3A, our results revealed that fluorescence intensity in the 37 • C group was significantly higher than that of the 25 • C group; however, there were no apparent differences when compared with the 4 • C group ( Figure 3B and Supplementary Figure S1D). These data suggest that Dot1l penetration may partially involve temperature-dependent cellular entry pathways, although some references in the field highlighted an energy-independent contribution to CPP uptake [29][30][31]. To further understand the precise mechanism of Dotl1 peptide uptake, selective and specific interventions in endocytosis studies were conducted.…”