Cellular import of synthetic peptide using a cell-permeable sequence in plant protoplasts |
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| Institution: | 1. Laboratoire de biotechnologie et amélioration des plantes, Inra (UA 3013), Institut national polytechnique-école nationale supérieure agronomique, pôle de biotechnologies végétales, chemin de Borde-Rouge, 31326 Castanet-Tolosan, France;2. Pôle de biotechnologies végétales, université Paul-Sabatier (CNRS, UMR 5546), 24, chemin de Borde-Rouge, BP 17 Auzeville, 31326 Castanet-Tolosan, France;3. Pôle de biotechnologies végétales (IFR40), 24 chemin de Borde-Rouge, BP 17 Auzeville, 31326 Castanet-Tolosan, France;1. Department of Zoology, MBB College, Tripura, India;2. Department of Zoology, Woman’s College, Tripura, India;3. School of Bioscience, IIT Kharagpur, Kharagpur, India;4. Department of Bio Engineering, National Institute of Technology, Agartala, Tripura, India;1. Textile Engineering Department, Functional Fibrous Structures & Environmental Enhancement (FFSEE), Amirkabir Nanotechnology Research Institute (ANTRI), Amirkabir University of Technology, Tehran, Iran;2. Skin Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran;3. Department of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran;1. School of Electronic Engineering, Xi''an University of Posts and Telecommunications, Xi’an 710121, China;2. School of Physics, Northwest University, Xi’an 710127, China;3. School of Science, Xi''an University of Posts and Telecommunications, Xi’an 710121, China;1. National Engineering Laboratory for Advanced Yarn and Fabric Formation and Clean Production, Wuhan Textile University, Wuhan 430073, PR China;2. Hubei Collaborative Innovation Center for Advanced Organic Chemical Materials & Key Laboratory for the Synthesis and Application of Organic Functional Molecules, Ministry of Education & College of Chemistry & Chemical Engineering, Hubei University, Wuhan 430062, PR China;3. Institute for Frontier Materials, Deakin University, Geelong, Victoria 3216, Australia;1. Hubei Key Laboratory of the Forensic Science, Hubei University of Police, Wuhan 430035, China;2. Hubei Collaborative Innovation Center for Advanced Organic Chemical Materials & Ministry-of-Education Key Laboratory for the Synthesis and Application of Organic Functional Molecules & College of Chemistry and Chemical Engineering, Hubei University, Wuhan 430062, China |
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| Abstract: | In this paper, we describe a rapid method to incorporate biologically active synthetic peptide in plant protoplasts. The peptides used contain a hydrophobic membrane permeable sequence as a carrier for the import through the plasma membrane. The membrane permeable sequence corresponds to the h-region, the more hydrophobic domain found in the signal peptide of secreted proteins. To evaluate the feasibility of the method, we synthesized a cell-permeable peptide with an h-region of a plant signal peptide plus residues 410–419 of the human c-myc oncogene product. Detection was performed via fluorescence analysis using specific monoclonal anti-c-myc primary antibody and FITC-conjugated secondary antibody. No saturation of import was observed, suggesting that the mechanisms involved do not require energy. The half-life time of the internalized peptide was estimated and results indicate that peptide concentration into protoplasts was constant for 8 h following incorporation. This method is complementary to microinjection or to the use of membrane permeabilizing reagents to study in vivo protein–protein or DNA–protein interactions. Finally, this method was used to analyse a putative interaction between the conserved cytoplasmic tail of a transmembrane receptor (HaELP, Helianthus annuus EGF receptor like protein) and the cytoskeleton. No interaction was found between these components. |
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