The basic domain of Rev from human immunodeficiency virus type 1 specifically blocks the entry of U4/U6.U5 small nuclear ribonucleoprotein in spliceosome assembly.
Human immunodeficiency virus type 1 (HIV-1) encodes a regulatory protein, Rev, which is required for cytoplasmic expression of incompletely spliced viral mRNA. Rev binds to a cis-acting Rev-responsive element (RRE) located within the env region of HIV-1. It has previously been shown that a 17-amino-acid peptide, corresponding to the basic domain of Rev, specifically inhibited in vitro the splicing of mRNAs containing the RRE. In this reaction, the peptide acts after an ATP-dependent step in the spliceosome assembly resulting in an accumulation of a 45-50S splicing-deficient complex. Characterization of this complex revealed that the basic domain of Rev does not interfere with U1 small nuclear ribonucleoprotein binding but blocks the entry of U4, U5, and U6 small nuclear RNAs into the spliceosome. Binding of U2 small nuclear ribonucleoprotein was partially inhibited. The critical nature of the oligomeric structure of RRE has been investigated both in vitro and in vivo. Reporter genes that contained one, three, or six repeated-monomer high-affinity Rev binding sites (IIB) within an intron yielded a correlation among the oligomeric state of bound Rev; inhibition of splicing; ability to block the assembly of U4, U5, and U6 small nuclear RNAs in the spliceosome in vitro; and level of Rev response in vivo.
Kjems
J
J
Department of Molecular Biology, University of Aarhus, Denmark.
Sharp
P A
PA
D001483:Base Sequence; D015152:Blotting, Northern; D015694:Gene Products, rev / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology*; D015752:Genes, env; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010446:Peptide Fragments / Q000494:pharmacology; D010957:Plasmids; D011485:Protein Binding; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D015183:Restriction Mapping; D017414:Ribonucleoprotein, U4-U6 Small Nuclear / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017381:Spliceosomes / Q000187:drug effects / Q000378:metabolism*; D054321:rev Gene Products, Human Immunodeficiency Virus
D015694:Gene Products, rev; D010446:Peptide Fragments; D012322:RNA Precursors; D012333:RNA, Messenger; D012342:RNA, Small Nuclear; D012367:RNA, Viral; D017414:Ribonucleoprotein, U4-U6 Small Nuclear; D054321:rev Gene Products, Human Immunodeficiency Virus