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- Juks, Carmen, et al.
(författare)
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The role of endocytosis in the uptake and intracellular trafficking of PepFect14-nucleic acid nanocomplexes via class A scavenger receptors
- 2015
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Ingår i: Biochimica et Biophysica Acta - Biomembranes. - : Elsevier BV. - 0005-2736 .- 1879-2642. ; 1848:12, s. 3205-3216
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Tidskriftsartikel (refereegranskat)abstract
- Cell penetrating peptides are efficient tools to deliver various bioactive cargos into cells, but their exactfunctioning mechanism is still debated. Recently, we showed that a delivery peptide PepFect14 condenses oligonudeotides (ON) into negatively charged nanocomplexes that are taken up by cells via class A scavenger receptors (SR-As). Here we unraveled the uptake mechanism and intracellular trafficking of PF14-ON nanocomplexes in HeLa cells. Macropinocytosis and caveolae-mediated endocytosis are responsible for the intracellular functionality of nucleic acids packed into nanocomplexes. However, only a negligible fraction of the complexes were trafficked to endoplasmic reticulum or Golgi apparatus the common destinations of caveolar endocytosis. Neither were the PF14-SCO nanocomplexes routed to endo-lysosomal pathway, and they stayed in vesicles with slightly acidic pH, which were not marked with LysoSensor. Naked ON, in contrary, was rapidly targeted to acidic vesicles and lysosomes. The transmission electron microscopy analysis of interactions between SR-As and PF14-ON nanocomplexes on ultrastructural level revealed that nanocomplexes localized on the plasma membrane in close proximity to SR-As and their colocalization is retained in cells, suggesting that PF14-ON complexes associate with targeted receptors.
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| 2. |
- Raeaegel, Helin, et al.
(författare)
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Cell-penetrating peptide secures an efficient endosomal escape of an intact cargo upon a brief photo-induction
- 2013
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Ingår i: Cellular and Molecular Life Sciences (CMLS). - : Springer Science and Business Media LLC. - 1420-682X .- 1420-9071. ; 70:24, s. 4825-4839
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Tidskriftsartikel (refereegranskat)abstract
- Since their discovery, cell-penetrating peptides (CPPs) have provided a novel, efficient, and non-invasive mode of transport for various (bioactive) cargos into cells. Despite the ever-growing number of successful implications of the CPP-mediated delivery, issues concerning their intracellular trafficking, significant targeting to degradative organelles, and limited endosomal escape are still hindering their widespread use. To overcome these obstacles, we have utilized a potent photo-induction technique with a fluorescently labeled protein cargo attached to an efficient CPP, TP10. In this study we have determined some key requirements behind this induced escape (e.g., dependence on peptide-to-cargo ratio, time and cargo), and have semi-quantitatively assessed the characteristics of the endosomes that become leaky upon this treatment. Furthermore, we provide evidence that the photo-released cargo remains intact and functional. Altogether, we can conclude that the photo-induced endosomes are specific large complexes-condensed non-acidic vesicles, where the released cargo remains in its native intact form. The latter was confirmed with tubulin as the cargo, which upon photo-induction was incorporated into microtubules. Because of this, we propose that combining the CPP-mediated delivery with photo-activation technique could provide a simple method for overcoming major limitations faced today and serve as a basis for enhanced delivery efficiency and a subsequent elevated cellular response of different bioactive cargo molecules.
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