Cationic antimicrobial peptides play important roles in innate immunity. Compared with extensive studies on
peptide-bacteria interactions, little is known about
peptide-human cell interactions. Using human cervical
carcinoma HeLa and fibroblastic TM12 cells, we investigated the cellular uptake of fluorescent analogues of the two representative
antimicrobial peptides magainin 2 and
buforin 2 in comparison with the representative Arg-rich cell-penetrating Tat-(47-57)
peptide (
YGRKKRRQRRR). The dose, time, temperature, and energy dependence of translocation suggested that the three
peptides cross cell membranes through different mechanisms. The
magainin peptide was internalized within a time scale of
tens of minutes. The cooperative concentration dependence of uptake suggested that the
peptide forms a pore as an intermediate similar to the observations in model membranes. Furthermore, the translocation was coupled with cytotoxicity, which was larger for
tumor HeLa cells. In contrast, the buforin
peptide translocated within 10 min by a temperature-independent, less concentration-dependent passive mechanism without showing any significant cytotoxicity at the highest concentration investigated (100 microm). The uptake of the Tat
peptide was proportional to the
peptide concentration, and the concentration dependence was lost upon
ATP depletion. The
peptide exhibited a moderate cytotoxicity at higher concentrations. The time course did not show saturation even after 120 min. The buforin
peptide, covalently attached to the 28-kDa
green fluorescent protein, also entered cells, suggesting a potency of the
peptide as a vector for macromolecular delivery into cells. However, the mechanism appeared to be different from that of the parent
peptide.