title: Relative contribution of photo-addition, helper oligonucleotide and RNase H to the antisense effect of psoralen-oligonucleotide conjugates, on in vitro translation of Leishmania mRNAs.
abstract: We investigated the properties of two antisense oligonucleotides, 11 alpha Pso and 14TMP, 11 and 14 nucleotides long, respectively, and conjugated to psoralen derivatives. These oligonucleotides were complementary to the mini-exon sequence of Leishmania amazonensis. Upon ultraviolet (UV) irradiation these oligomers were selectively cross-linked to DNA or RNA target sequences, either 14 or 35 nucleotides long. The yield of photo-addition was much lower on the longer targets than on the shorter ones, due to the presence of a hairpin structure. The co-addition of a helper oligonucleotide, whose binding site, on the 35-mer, was adjacent to that of the psoralen-derivatized antisense oligomer, improved the cross-linking efficiency. We then determined the effect of 14TMP on in vitro translation of Leishmania mRNA in cell-free extracts. Non-irradiated antisense oligonucleotide/mRNA complexes reduced the protein synthesis in wheat germ extract but not in rabbit reticulocyte lysate. Conversely, UV irradiation induced a 14TMP-dependent reduction of translation in reticulocyte lysate whereas the inhibition was not improved in the wheat germ extract. These results are discussed with respect to the involvement of RNase-H in the oligonucleotide-mediated effect on protein synthesis.
authors.affiliation: Laboratoire de Biophysique Moléculaire, INSERM U386, Université de Bordeaux II, France.
authors.forename: E
authors.initials: E
authors.lastname: Pascolo
authors.forename: D
authors.initials: D
authors.lastname: Hudrisier
authors.forename: B
authors.initials: B
authors.lastname: Sproat
authors.forename: T T
authors.initials: TT
authors.lastname: Nguyen
authors.forename: J J
authors.initials: JJ
authors.lastname: Toulmé
mesh_terms: D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D005091:Exons / Q000235:genetics; D011564:Furocoumarins / Q000494:pharmacology*; D007891:Leishmania / Q000235:genetics*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009843:Oligoribonucleotides / Q000494:pharmacology*; D017319:Photosensitizing Agents / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects* / Q000528:radiation effects; D016372:RNA, Antisense / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D016914:Ribonuclease H / Q000494:pharmacology; D014466:Ultraviolet Rays
chemical_list: D011564:Furocoumarins; D009843:Oligoribonucleotides; D017319:Photosensitizing Agents; D016372:RNA, Antisense; D012333:RNA, Messenger; D016914:Ribonuclease H