Arginine-rich cross-linking peptides with different SV40 nuclear localization signal content as vectors for intranuclear DNA delivery

Abstract

The major barriers for intracellular DNA transportation by cationic polymers are their toxicity, poor endosomal escape and inefficient nuclear uptake. Therefore, we designed novel modular peptide-based carriers modified with SV40 nuclear localization signal (NLS). Core peptide consists of arginine, histidine and cysteine residues for DNA condensation, endosomal escape promotion and interpeptide cross-linking, respectively. We investigated three polyplexes with different NLS content (10 mol%, 50 mol% and 90 mol% of SV40 NLS) as vectors for intranuclear DNA delivery. All carriers tested were able to condense DNA, to protect it from DNAase I and were not toxic to the cells. We observed that cell cycle arrest by hydroxyurea did not affect transfection efficacy of NLS-modified carriers which we confirmed using quantitative confocal microscopy analysis. Overall, peptide carrier modified with 90 mol% of SV40 NLS provided efficient transfection and nuclear uptake in non-dividing cells. Thus, incorporation of NLS into arginine-rich cross-linking peptides is an adequate approach to the development of efficient intranuclear gene delivery vehicles.

DNA Vaccines—How Far From Clinical Use?

“…As a consequence, in some approaches the SV40 enhancer site was included into the vector backbone [ 11 ]. Alternatively, the NLS activity of virus-derived peptides like the SV40 large T-antigen was exploited to facilitate nuclear plasmid translocation by attaching these peptides either directly to the vector backbone [ 119 ] or to DNA delivery systems [ 120 ].…”

Mitochondria and neuroprotection in stroke: Cationic arginine-rich peptides (CARPs) as a novel class of mitochondria-targeted neuroprotective therapeutics

“…in heparin sulphate proteoglycans), phosphate (e.g. in phospholipid head groups) and carboxylate (in phospholipid phosphatidylserine and protein receptors) moieties present on the plasma membrane or organelle membrane structures, such as those present in mitochondria, Golgi apparatus, endoplasmic reticulum, or the nucleus [170][171][172][173][174]. The electrostatic interactions between arginine residues and anionic structures present on membranes is a critical requirement for CARPs to cross typically impervious membranes,…”

Lipoaminoacids Enzyme-Based Production and Application as Gene Delivery Vectors

“…The toxicity, poor endosomal escape, and inefficient nuclear uptake of cationic polymers, which constitute major barriers for DNA delivery, can be overcome by cationic LAAs due to their improved biocompatibility, low cytotoxicity, and pH-dependent behavior that promotes structural changes from lamellar to inverted hexagonal phases at the acidic endosomal compartment facilitating DNA release [7][8][9]16,96]. Recently, intranuclear DNA delivery by cationic LAAs was further improved while using arginine-rich cross-linking peptides with different SV40 nuclear localization signal (NLS) content [103]. The nuclear import pathway is the classical transport between the cytoplasm and the nucleus, and the Arg-rich peptides modified with 90 mol% of SV40 NLS provided efficient intranuclear DNA delivery vehicles [103].…”

“…Recently, intranuclear DNA delivery by cationic LAAs was further improved while using arginine-rich cross-linking peptides with different SV40 nuclear localization signal (NLS) content [103]. The nuclear import pathway is the classical transport between the cytoplasm and the nucleus, and the Arg-rich peptides modified with 90 mol% of SV40 NLS provided efficient intranuclear DNA delivery vehicles [103]. Thus, LAAs are efficient and safer potential alternatives to both conventional cationic surfactants and cationic polymers as non-viral gene delivery vectors.…”