Presence of negative and positive cis-acting RNA splicing elements within and flanking the first tat coding exon of human immunodeficiency virus type 1.
The human immunodeficiency virus type 1 (HIV-1) RNA follows a complex splicing pathway in which a single primary transcript either remains unspliced or is alternatively spliced to more than 30 different singly and multiply spliced mRNAs. We have used an in vitro splicing assay to identify cis elements within the viral genome that regulate HIV-1 RNA splicing. A novel splicing regulatory element (SRE) within the first tat coding exon has been detected. This element specifically inhibits splicing at the upstream 3' splice site flanking this tat exon. The element only functions when in the sense orientation and is position dependent when inserted downstream of a heterologous 3' splice site. In vivo, an HIV-1 SRE mutant demonstrated a decrease in unspliced viral RNA, increased levels of single- and double-spliced tat mRNA, and reduced levels of env and rev mRNAs. In addition to the negative cis-acting SRE, the flanking 5' splice site downstream of the first tat coding exon acts positively to increase splicing at the upstream 3' splice sites. These results are consistent with hypotheses of bridging interactions between cellular factors that bind to the 5' splice site and those that bind at the upstream 3' splice site.
Amendt
B A
BA
Department of Microbiology, University of Iowa, Iowa City 52242.
Hesslein
D
D
Chang
L J
LJ
Stoltzfus
C M
CM
D017398:Alternative Splicing; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers; D005091:Exons; D015967:Gene Expression Regulation, Viral; D015756:Genes, tat; D015497:HIV-1 / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D012333:RNA, Messenger / Q000096:biosynthesis*; D012045:Regulatory Sequences, Nucleic Acid; D015183:Restriction Mapping; D014162:Transfection
D017931:DNA Primers; D012333:RNA, Messenger