The influence of alkyl pyridinium sponge toxins on membrane properties, cytotoxicity, transfection and protein expression in mammalian cells.
The ability of two alkyl pyridinium sponge toxin preparations (poly-APS and halitoxin) to form transient pores/lesions in cell membranes and allow transfection of plasmid cDNA have been investigated using HEK 293 cells. Poly-APS and halitoxin preparations caused a collapse in membrane potential, reductions in input resistance and increased Ca2+ permeability. At least partial recovery was observed after poly-APS application but recovery was more rarely seen with halitoxin. The transfection with plasmid cDNAs for an enhanced green fluorescent protein (EGFP) and human tumour necrosis factor receptor 2 (TNFR2) was assessed for both toxin preparations and compared with lipofectamine. Stable transfection was achieved with poly-APS although it was less efficient than lipofectamine. These results show that viable cells transfected with alien cDNA can be obtained using novel transient pore-forming alkyl pyridinium sponge toxins and a simple pre-incubation protocol. This provides the first proof of principle that pore-forming alkyl pyridinium compounds can be used to deliver cDNA to the intracellular environment without permanently compromising the plasma membrane.
Tucker
Steven J
SJ
Department of Biomedical Sciences, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, Scotland AB25 2ZD, UK. bms197@abdn.ac.uk
McClelland
David
D
Jaspars
Marcel
M
Sepcić
Kristina
K
MacEwan
David J
DJ
Scott
Roderick H
RH
D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis / Q000235:genetics; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004705:Endocytosis; D049452:Green Fluorescent Proteins; D006801:Humans; D008055:Lipids; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008387:Marine Toxins / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects; D018613:Microscopy, Confocal; D010957:Plasmids; D011108:Polymers / Q000494:pharmacology*; D011161:Porifera; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D011726:Pyridinium Compounds / Q000494:pharmacology*; D018124:Receptors, Tumor Necrosis Factor / Q000096:biosynthesis / Q000235:genetics; D047889:Receptors, Tumor Necrosis Factor, Type II; D014162:Transfection / Q000379:methods
D015703:Antigens, CD; D018076:DNA, Complementary; D008055:Lipids; C086724:Lipofectamine; D008164:Luminescent Proteins; D008387:Marine Toxins; D011108:Polymers; D011506:Proteins; D011726:Pyridinium Compounds; D018124:Receptors, Tumor Necrosis Factor; D047889:Receptors, Tumor Necrosis Factor, Type II; C112488:halitoxin; C115254:poly-APS; D049452:Green Fluorescent Proteins