Probing alternative foldings of the HIV‐1 leader RNA by antisense oligonucleotide scanning arrays

Scanning arrays of antisense DNA oligonucleotides provide a novel and systematic means to study structural features within an RNA molecule. We used this approach to probe the structure of the untranslated leader of the human immunodeficiency virus type 1 (HIV‐1) RNA genome. This 335 nt RNA encodes m...

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Bibliographic Details
Published in:Nucleic acids research 2004-01, Vol.32 (2), p.819-827
Main Authors: Ooms, Marcel, Verhoef, Koen, Southern, Edwin, Huthoff, Hendrik, Berkhout, Ben
Format: Article
Language:English
Subjects:
Base Pairing
Base Sequence
Electrophoretic Mobility Shift Assay
HIV Long Terminal Repeat - genetics
HIV-1 - genetics
Human immunodeficiency virus
Molecular Sequence Data
Nucleic Acid Conformation
Nucleic Acid Hybridization
Oligonucleotide Array Sequence Analysis
Oligonucleotides, Antisense - chemistry
Oligonucleotides, Antisense - genetics
Oligonucleotides, Antisense - metabolism
RNA, Viral - chemistry
RNA, Viral - genetics
RNA, Viral - metabolism
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Summary:Scanning arrays of antisense DNA oligonucleotides provide a novel and systematic means to study structural features within an RNA molecule. We used this approach to probe the structure of the untranslated leader of the human immunodeficiency virus type 1 (HIV‐1) RNA genome. This 335 nt RNA encodes multiple important replication signals and adopts two mutually exclusive conformations. The poly(A) and the dimer initiation signal (DIS) sequences of the leader RNA are base‐paired in the long‐distance interaction (LDI) conformation, but both domains form distinct hairpins in the branched multiple hairpins (BMH) conformation. An RNA switch mechanism has been proposed to regulate the activity of the DIS dimerization signal that is masked in one, yet exposed in the other conformation. The two RNA conformations demonstrate discrete differences in the array‐based hybridization patterns. LDI shows increased hybridization in the poly(A) region and decreased hybridization in the DIS region when compared with BMH. These results provide additional evidence for the structure models of the two alternative leader RNA conformations. We also found a correlation between the efficiency of oligonucleotide hybridization and the accessibility of the RNA structure as determined by chemical and enzymatic probing in previous studies. The array approach therefore provides a very sensitive method to detect structural differences in related transcripts.
ISSN:0305-1048
1362-4962
1362-4962
DOI:10.1093/nar/gkh206