Uptake and cellular distribution of nucleolar targeting peptides (NrTPs) in different cell types

Nucleolar targeting peptides (NrTPs) are a family of cell penetrating peptides (CPPs) derived from crotamine, a rattlesnake venom toxin. They were named NrTPs for their remarkable nucleolus-homing properties and have been studied for their potential as drug delivery vehicles. Live cell microscopy experiments were conducted to monitor NrTP uptake and distribution in different cell types, including primary cells (PBMCs and erythrocytes) and different immortalized cell lines (HeLa, BHK21, BV-173, and MOLT-4). Uptake dependence on cell type (primary vs. immortalized, suspension vs. adherent, cancer vs. healthy cells), peptide concentration and cell viability were evaluated. To gain further insight on the internalization mechanism, uptake kinetics was also monitored. Results showed the uptake and distribution pattern as strongly dependent on peptide sequence, peptide concentration and membrane constituents. Under similar conditions, NrTP6 is more internalized than NrTP1, NrTP2 and NrTP5. Additionally, while internalization of NrTP7 and NrTP8 may cause cytotoxicity, NrTP6 is noncytotoxic. Higher peptide concentrations can be correlated to nucleolar targeting, although even at low concentrations a residual number of cells reveal positive nucleolar labeling. NrTPs were successfully internalized into all cell types tested except erythrocytes.