RESUMEN
Plasma membranes of animal cells are generally impermeable to macromolecules. Protein transduction mediated by protein transduction domains (PTDs) covalently cross-linked to cargoes for cellular internalization has previously been demonstrated. Peptides with PTDs could be an effective way to deliver proteins into living cells or tissues in vitro. In this report, we demonstrate that arginine-rich intracellular delivery (AID) peptides are able to facilitate the delivery of proteins into animal cells and to penetrate skin tissues rapidly. This cellular internalization and transdermal delivery of proteins is mediated by non-toxic AID peptides in a non-fusion protein and non-conjugation dependent manner. The efficiency of intracellular transport is further increased in the presence of chemical enhancer oleic acid. The mechanism of the AID-mediated cellular entry may involve macropinocytosis and actin rearrangement. Thus, we confirm that direct delivery of bioactive proteins into living cells and tissues mediated by non-covalent actions of AID peptides represents a useful strategy in pharmaceutics, therapeutics and cosmetics.
Asunto(s)
Arginina , Membrana Celular/metabolismo , Proteínas Fluorescentes Verdes/metabolismo , Péptidos , Administración Cutánea , Animales , Línea Celular Tumoral , Portadores de Fármacos , Proteínas Fluorescentes Verdes/administración & dosificación , Humanos , Ratones , Plásmidos , Transporte de ProteínasRESUMEN
RNA interference has recently become a useful research tool for the studies of gene functions, regulations, and therapies. The double-stranded RNA is utilized to induce the sequence-specific gene silencing. To achieve this goal of specific gene silencing, a proper delivery system of siRNA is highly demanded. A number of approaches for delivering siRNA have been explored over the last few years. In the present study, we demonstrated a simple peptide-based siRNA delivery system in mammalian cells. A GC-EGFP cell line stably expressing enhanced green fluorescent protein was established from stable transfection of human gastric carcinoma cells. The synthetic nona-arginine peptide, an arginine-rich intracellular delivery peptide, or called protein transduction domain peptide, could noncovalently form stable complexes with EGFP siRNA and deliver these mixtures into cells. After entry, siRNA appeared to stay in perinuclear regions within cell, and ultimately fulfilled its targeted egfp gene silencing. These data were in consonance with that RNA-induced silencing complex components could be also localized to these perinuclear regions, creating a focal point for RNA interference factories. In the future, this non-toxic peptide may be proved to be a useful tool for the delivery of exogenous siRNA in RNA interference research.
Asunto(s)
Arginina , Silenciador del Gen , Proteínas Fluorescentes Verdes/metabolismo , Péptidos , Interferencia de ARN , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Animales , Transporte Biológico , Caenorhabditis elegans/genética , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , TransfecciónRESUMEN
Plasma membranes of plant or animal cells are generally impermeable to peptides or proteins. Many basic peptides have previously been investigated and covalently cross-linked with cargoes for cellular internalization. In the current study, we demonstrate that arginine-rich intracellular delivery (AID) peptides are able to deliver fluorescent proteins or beta-galactosidase enzyme into animal and plant cells, as well as animal tissue. Cellular internalization and transdermal delivery of protein could be mediated by effective and nontoxic AID peptides in a neither fusion protein nor conjugation fashion. Therefore, noncovalent AID peptides may provide a useful strategy to have active proteins function in living cells and tissues in vivo.