title: Use of a nonviral vector to express a chimeric tRNA-ribozyme against lymphocytic choriomeningitis virus: cytoplasmic accumulation of a catalytically competent transcript but minimal antiviral effect.
abstract: RNA polymerase III promoters direct the ubiquitous, high-level, expression of small, stable RNAs such as tRNAs, and thus are attractive candidates for achieving stable expression of small therapeutic (e.g., antiviral) molecules, such as ribozymes or antisense RNAs. In this article, we describe the use of a nonviral vector containing a tRNA promoter to express an antilymphocytic choriomeningitis virus (LCMV) ribozyme (tRNA-Rib5). The chimeric tRNA-ribozyme is specifically and efficiently transcribed by pol III in cell-free extracts, and the resulting transcript has appropriate ribozyme activity. In tissue culture studies, high levels of chimeric transcripts were readily detectable and were transported to the cytoplasm, the site of LCMV replication. Despite accumulation of tRNA-Rib5 in the cytoplasm of stably transformed cell clones, antiviral effects were minimal or absent. The implications of these findings and the potential use of this vector system for in vivo studies requiring the delivery of small molecules are discussed.
authors.affiliation: Department of Neuropharmacology, Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: J R
authors.initials: JR
authors.lastname: Gebhard
authors.forename: C M
authors.initials: CM
authors.lastname: Perry
authors.forename: S
authors.initials: S
authors.lastname: Mahadeviah
authors.forename: J L
authors.initials: JL
authors.lastname: Whitton
mesh_terms: D016475:3T3 Cells; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology* / Q000627:therapeutic use; D002678:Chimera; D002522:Chlorocebus aethiops; D002999:Clone Cells; D003593:Cytoplasm / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D005822:Genetic Vectors / Q000378:metabolism* / Q000494:pharmacology / Q000627:therapeutic use; D008217:Lymphocytic choriomeningitis virus / Q000187:drug effects* / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000494:pharmacology / Q000627:therapeutic use; D010957:Plasmids / Q000235:genetics / Q000378:metabolism* / Q000627:therapeutic use; D011401:Promoter Regions, Genetic / Q000235:genetics; D012320:RNA Polymerase III / Q000187:drug effects; D016337:RNA, Catalytic / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology / Q000627:therapeutic use; D012358:RNA, Transfer, Met / Q000096:biosynthesis* / Q000235:genetics / Q000627:therapeutic use; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics; D014709:Vero Cells
chemical_list: D000998:Antiviral Agents; D016376:Oligonucleotides, Antisense; D016337:RNA, Catalytic; D012358:RNA, Transfer, Met; D012320:RNA Polymerase III