tailieunhanh - Báo cáo khoa học: Study of uptake of cell penetrating peptides and their cargoes in permeabilized wheat immature embryos

The uptake of five fluorescein labeled cell-penetrating peptides (Tat, Tat2, mutated-Tat, peptide vascular endothelial-cadherin and transportan) was studied in wheat immature embryos. Interestingly, permeabilization treat-ment of the embryos with toluene⁄ethanol (1 : 20, v⁄v with permeabiliza-tion buffer) resulted in a remarkably higher uptake of cell-penetrating peptides, whereas nonpermeabilized embryos failed to show significant cell-penetrating peptide uptake, as observed under fluorescence microscope and by fluorimetric analysis | ỊFEBS Journal Study of uptake of cell penetrating peptides and their cargoes in permeabilized wheat immature embryos Archana Chugh and Francois Eudes Lethbridge Research Centre Agriculture and Agri-Food Canada Alberta Canada Keywords cell membrane permeabilization cell-penetrating peptide endocytosis macropinocytosis nanocarrier Correspondence F. Eudes Lethbridge Research Centre Agriculture and Agri-Food Canada PO Box 3000 5403 1st Avenue South Lethbridge Alberta T1J 4B1 Canada Fax 1 403 382 3156 Tel 1 403 317 3338 E-mail eudesf@ Received 16 November 2007 revised 8 February 2008 accepted 3 March 2008 doi The uptake of five fluorescein labeled cell-penetrating peptides Tat Tat2 mutated-Tat peptide vascular endothelial-cadherin and transportan was studied in wheat immature embryos. Interestingly permeabilization treatment of the embryos with toluene ethanol 1 20 v v with permeabiliza-tion buffer resulted in a remarkably higher uptake of cell-penetrating peptides whereas nonpermeabilized embryos failed to show significant cellpenetrating peptide uptake as observed under fluorescence microscope and by fluorimetric analysis. Among the cell-penetrating peptides investigated Tat monomer Tat showed highest fluorescence uptake greater in permeabilized embryos than the nonpermeabilized embryos. On the other hand mutated-Tat serving as negative control did not show comparable fluorescence levels even in permeabilized embryos. A glucuronidase histochemical assay revealed that Tat peptides can efficiently deliver functionally active b-glucuronidase GUS enzyme in permeabilized immature embryos. Tat2-mediated GUS enzyme delivery showed the highest number of embryos with GUS uptake upon permeabilization treatment with toluene ethanol 1 40 v v with permeabilization buffer whereas only of nonpermeabilized embryos showed Tat2-mediated GUS uptake. Low temperature endocytosis and macropinocytosis inhibitors reduced delivery .