Efficacy of an amphipathic oligopeptide to shuttle and release a cis-acting DNA decoy into human cells.
We investigated the ability of an amphipathic oligopeptide to carry a synthetic dsDNA oligonucleotide inside human cells. The oligonucleotide was designed as a decoy binding site for the transcriptional activator of the methylguanine-DNA methyltransferase (MGMT) gene. The complex oligopeptide and decoy were administered to MCF10A exponentially growing cells, and the uptake was monitored by flow cytometry. After a 1-h exposure, almost all of the MCF10A cells were fluorescent, indicating that all of the cells had been transfected. By increasing the time, the fluorescence intensity per cell rapidly increased to a plateau at the 8-h time point. RT-PCR analysis of the MGMT gene was used as the molecular readout of the intracellular activity of the DNA decoy. MCF10A cells transfected with the oligopeptide/decoy complex showed a strong reduction in MGMT mRNA. Here, we discuss the advantages of using amphipathic oligopeptides as carriers of short DNA sequences.
Citti
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Istituto di Mutagenesi e Differenziamento, CNR, Pisa, Italy.
Rovero
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P
Colombo
M G
MG
Mariani
L
L
Poliseno
L
L
Rainaldi
G
G
D001483:Base Sequence; D002455:Cell Division; D002460:Cell Line; D004247:DNA / Q000008:administration & dosage* / Q000737:chemistry; D017931:DNA Primers; D004337:Drug Carriers; D004587:Electrophoresis, Agar Gel; D005434:Flow Cytometry; D006801:Humans; D009842:Oligopeptides / Q000008:administration & dosage*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D017931:DNA Primers; D004337:Drug Carriers; D009842:Oligopeptides; D004247:DNA