Document 0393
 DOCN  M9460393
 TI    Binding of nuclear proteins to HTLV-II cis-acting repressive sequence
       (CRS) RNA correlates with CRS function.
 DT    9404
 AU    Black AC; Ruland CT; Luo J; Bakker A; Fraser JK; Rosenblatt JD;
       Department of Medicine, UCLA School of Medicine 90024.
 SO    Virology. 1994 Apr;200(1):29-41. Unique Identifier : AIDSLINE
       MED/94174729
 AB    The shift from viral regulatory to structural gene expression in human
       T-cell leukemia virus types I (HTLV-I) and II (HTLV-II) is mediated by
       Rex. We have previously shown that HTLV-II Rex acts through an element
       in R/U5 of the 5' long terminal repeat (LTR), the Rex-responsive element
       (RxRE), and that Rex protein binds to specific RNA sequences, the Rex
       binding element (RBE), contained within the RxRE (Black et al., J.
       Virol. 65, 6645-6653, 1991b). Rex action through the RBE (nt 405-520)
       overcomes the inhibition of expression conferred by a contiguous LTR RNA
       regulatory element, which contains cis-acting repressive sequences (CRS;
       nt 520-630) that are not bound by Rex protein (Black et al., Virology,
       181, 433-444, 1991a). We now show by electrophoretic mobility shift
       assay (EMSA) that cellular proteins in a HeLa nuclear extract bind
       specifically to RNA transcripts containing the HTLV-II CRS. Using
       ultraviolet (uv) crosslinking of gel-retarded bands, we identified a
       major protein species of approximately 60 kDa, p60CRS, that binds to CRS
       RNA and, with weaker affinity, to RBE RNA. In addition, a distinct
       40-kDa protein, p40CRS, binds to U5 RNA (nt 645-750) downstream from the
       CRS. Specific deletions within CRS RNA can reduce or abrogate binding to
       this 60-kDa protein. EMSA and uv crosslinking assays also suggest that
       both p60CRS and p40CRS interact with CRS RNA. CRS function in a 5'
       LTR-linked gene expression assay correlates with the ability of both
       p60CRS and p40CRS to interact with 5' LTR RNA in vitro.
 DE    Base Sequence  Chloramphenicol Acetyltransferase/GENETICS  Cross-Linking
       Reagents  *Gene Expression Regulation, Viral  Hela Cells  Human
       HTLV-II/*GENETICS  Molecular Sequence Data  Nuclear Proteins/*METABOLISM
       Protein Binding  Recombinant Fusion Proteins/METABOLISM  Repetitive
       Sequences, Nucleic Acid/*GENETICS  Repressor Proteins/*METABOLISM  RNA,
       Viral/GENETICS/*METABOLISM  Support, U.S. Gov't, P.H.S.  Transfection
       Ultraviolet Rays  JOURNAL ARTICLE

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