Inhibition of in vitro and ex vivo translation by a transplatin-modified oligo(2'-O-methylribonucleotide) directed against the HIV-1 gag-pol frameshift signal.
A 2'-O-methylribooligonucleotide containing a G1.U.G3 triad modified by trans-diamminedichloro-platinum(II) was targeted to the RNA region responsible for the gag-pol frameshifting during translation of the HIV-1 mRNA. The binding of the platinated oligonucleotide to its target RNA induced a rearrangement of the (G1, G3)-intrastrand crosslink, leading to the formation of an intermolecular oligonucleotide-RNA G-A crosslink. This resulted in the selective arrest of translation of a luciferase gene placed downstream of the HIV-1 frameshift signal both in a cell-free extract (rabbit reticulocyte lysate) and in RNA-transfected cells. A specific inhibition of luciferase activity was still observed when the oligonucleotide-RNA complex was not pre-formed prior to either translation or transfection. Moreover, a selective inhibition was also observed when the oligonucleotide and the plasmid DNA encoding the luciferase and bearing the RNA gag- pol frameshifting signal were co-transfected in NIH 3T3 cultured cells. Therefore the intra-strand-->interstrand conversion of the platinum crosslink kinetically competes with the translation machinery and blocks the polypeptide elongation. These transplatin-modified oligonucleotides which operate within a live cell on a 'real-time' basis and do not need an external triggering signal constitute a promising new class of selective reactive probes.
Aupeix-Scheidler
K
K
INSERM U.386, IFR Pathologies Infectieuses, Université Victor Segalen Bordeaux 2, 146 Rue Léo Saignat, 33076 Bordeaux Cedex, France,
Chabas
S
S
Bidou
L
L
Rousset
J P
JP
Leng
M
M
Toulmé
J J
JJ
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002945:Cisplatin / Q000737:chemistry*; D018965:Frameshifting, Ribosomal; D015685:Fusion Proteins, gag-pol / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009843:Oligoribonucleotides / Q000737:chemistry / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects*; D012367:RNA, Viral / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects
D015685:Fusion Proteins, gag-pol; D009843:Oligoribonucleotides; D012367:RNA, Viral; C034697:transplatin; D002945:Cisplatin