Dataset Information

Targeted Gene Delivery into Various Plastids Mediated by Clustered Cell-Penetrating and Chloroplast-Targeting Peptides.

ABSTRACT: The plastid is an organelle that functions as a cell factory to supply food and oxygen to the plant cell and is therefore a potential target for genetic engineering to acquire plants with novel photosynthetic traits or the ability to produce valuable biomolecules. Conventional plastid genome engineering technologies are laborious for the preparation of plant material, require expensive experimental instruments, and are time consuming for obtaining a transplastomic plant line that produces significant levels of the biomolecule of interest. Herein, a transient plastid transformation technique is presented using a peptide-based gene carrier. By formulating peptide/plasmid DNA complexes that combine the functions of both a cell-penetrating peptide and a chloroplast-targeting peptide, DNA molecules are translocated across the plant cell membrane and delivered to the plastid efficiently via vesicle formation and intracellular vesicle trafficking. A simple infiltration method enables the introduction of a complex solution into intact plants, and plastid-localized transgene expression is expeditiously observed in various types of plastids in differentiated cell types of several plants. The gene delivery technology thus provides a useful tool to rapidly engineer plastids in crop species.

SUBMITTER: Thagun C

PROVIDER: S-EPMC6891901 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Targeted Gene Delivery into Various Plastids Mediated by Clustered Cell-Penetrating and Chloroplast-Targeting Peptides.

Thagun Chonprakun C Chuah Jo-Ann JA Numata Keiji K

Advanced science (Weinheim, Baden-Wurttemberg, Germany) 20191023 23

The plastid is an organelle that functions as a cell factory to supply food and oxygen to the plant cell and is therefore a potential target for genetic engineering to acquire plants with novel photosynthetic traits or the ability to produce valuable biomolecules. Conventional plastid genome engineering technologies are laborious for the preparation of plant material, require expensive experimental instruments, and are time consuming for obtaining a transplastomic plant line that produces signif ...[more]

PMID: 31832328

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Project description:Many neurodegenerative disorders (NDDs) are characterized by aggregation of aberrant proteins and extensive oxidative stress in brain cells. As a treatment option for NDDs, RNA interference (RNAi) is a promising approach to suppress the activation of abnormal genes and negative regulators of antioxidant genes. Efficient neuro-targeted siRNA delivery requires a delicate optimization of nucleic acid carriers, quite distinct from putative pDNA carriers in regard to stable condensation and serum protection of siRNA, blood-brain barrier (BBB) bypass, effective siRNA delivery to brain cells, and functional release of bioactive siRNA at therapeutic levels. Here, we propose that a myristic acid conjugated, cell-penetrating peptide (transportan; TP), equipped with a transferrin receptor-targeting peptide (myr-TP-Tf), will lead to stable encapsulation of siRNA and targeted delivery of siRNA to brain cells overcoming the BBB. Myr-TP-Tf was successfully prepared by solid-phase peptide synthesis with high purity. Myr-TP-Tf-siRNA complexes formulated at a 20:1 (peptide-siRNA) molar ratio provided prolonged siRNA stability against serum and ribonuclease treatment. Fluorescence images clearly indicated that siRNA uptake was successfully achieved by myr-TP-Tf complexes in both a murine brain endothelioma and a human glioma cell line. The luciferase assay and the human placental alkaline phosphatase (hPAP) reporter assay results demonstrated the functional gene silencing effect of myr-TP-Tf-siRNA complexes in a human glioma cell line as well as in primary murine neurons/astrocytes, supportive of successful release of bioactive siRNA into the cytosol. Finally, the transcytosis assay revealed that favorable siRNA transport via receptor-mediated transcytosis was mediated by myr-TP-Tf complexes. In summary, these data suggest that myr-TP-Tf peptides possess promising properties as a vehicle for neuro-targeted siRNA delivery. We will further study this peptide in vitro and in vivo for transport mechanism kinetics and to validate its capability to deliver siRNA to the brain, respectively.

Dataset Information

Targeted Gene Delivery into Various Plastids Mediated by Clustered Cell-Penetrating and Chloroplast-Targeting Peptides.

Publications

Targeted Gene Delivery into Various Plastids Mediated by Clustered Cell-Penetrating and Chloroplast-Targeting Peptides.

Similar Datasets

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