Cellular activity of Rev response element RNA targeting metallopeptides.
The cellular chemistry of metallopeptide complexes designed to target and inactivate an HIV Rev response element (RRE) RNA sequence in vivo has been evaluated by use of an efficient cellular fluorescence assay. Transcribed messenger RNA encoding the green fluorescent protein (GFP) that includes a target RNA sequence is sensitive to cleavage chemistry mediated by metal derivatives of GGH(G)(x)TRQARRNRR RRWRERQR (x = 0, 1, 2, 4, 6). This results in a significant decrease in expression of GFP that can be quantified by fluorimetry. Optimal inactivation of the target RRE RNA was achieved with linkers where x = 0 or 1. Neither the Rev control peptide (lacking metal-binding or linker sequences) nor the metal-binding motif alone had any significant effect. Consequently, both the cleavage motif and the RNA targeting motif are essential to promote cellular cleavage of the target RRE RNA. However, target inactivation was also observed in experiments with metal-free peptide, consistent with recruitment of intracellular metal ion by the peptide following cellular uptake, with subsequent cleavage of the RRE target RNA. The RRE RNA cleavage activities of metallopeptide complexes were further confirmed by in vitro experiments and mammalian cell assays.
Jin
Yan
Y
Evans Laboratory of Chemistry, Ohio State University, 100 West 18th Avenue, Columbus, OH 43210, USA.
Cowan
J A
JA
D000465:Algorithms; D000595:Amino Acid Sequence; D002151:Calorimetry; D002470:Cell Survival; D003300:Copper / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D015195:Drug Design; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism; D015694:Gene Products, rev / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006678:HIV / Q000235:genetics*; D006801:Humans; D019169:Jurkat Cells; D008667:Metalloproteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009532:Nickel / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism*; D054321:rev Gene Products, Human Immunodeficiency Virus
D015694:Gene Products, rev; D008667:Metalloproteins; D012367:RNA, Viral; D054321:rev Gene Products, Human Immunodeficiency Virus; D049452:Green Fluorescent Proteins; D003300:Copper; D009532:Nickel