Nuclear targeting peptide scaffolds for lipofection of nondividing mammalian
cells
Ajit Subramanian, P. Ranganathan
& Scott L. Diamond
Institute for Medicine and Engineering, Department
of Chemical Engineering, 1010 Vagelos Research Laboratories, University of
Pennsylvania, Philadelphia, PA 19104.
Correspondence should be addressed to Scott L. Diamond sld@seas.upenn.eduGene transferendotheliumlipofectionnuclear import
Lipofection of nondividing cells is inefficient because much of the transfected
DNA is retained in endosomes, and that which escapes to the cytoplasm enters
the nucleus at low rates. To improve the final rate-limiting step of nuclear
import, we conjugated a nonclassical nuclear localization signal (NLS) containing
the M9 sequence of heterogeneous nuclear ribonucleoprotein (hnRNP) A1, to
a cationic peptide scaffold derived from a scrambled sequence of the SV40
T-antigen consensus NLS (ScT). The ScT was added to improve DNA binding of
the M9 sequence. Lipofection of confluent endothelium with plasmid complexed
with the M9−ScT conjugate resulted in 83% transfection and a 63-fold
increase in marker gene expression. The M9−ScT conjugate localized fluorescent
plasmid into the nucleus of permeabilized cells, and addition of the nuclear
pore blocker wheat germ agglutinin prevented nuclear import. This method of
gene transfer may lead to viral- and lipid-free transfection of nondividing
cells.
