SR proteins and hnRNP H regulate the splicing of the HIV-1 tev-specific exon 6D.
A naturally arising point mutation in the env gene of HIV-1 activates the aberrant inclusion of the cryptic exon 6D into most viral messages, leading to inefficient viral replication. We set out to understand how a single nucleotide substitution could cause such a dramatic change in splicing. We have determined that the exon 6D mutation promotes binding of the SR protein SC35 to the exon. Mutant exon 6D sequences function as a splicing enhancer when inserted into an enhancer-dependent splicing construct. hnRNP H family proteins bind to the enhancer as well; their binding is dependent on the sequence GGGA located just downstream of the point mutation and depletion-- reconstitution studies show that hnRNP H is essential for enhancer activity. A polypurine sequence located further downstream in exon 6D binds SR proteins but acts as an exonic splicing silencer. hnRNP H is required for interaction of U1 snRNP with the enhancer, independent of the point mutation. We propose that SC35 binding to the point mutation region may convert the hnRNP H-U1 snRNP complex into a splicing enhancer.
Caputi
Massimo
M
Department of Molecular, Cell and Developmental Biology, Sinsheimer Laboratory, University of California Santa Cruz, Santa Cruz, CA 95064, USA.
Zahler
Alan M
AM
D001483:Base Sequence; D004742:Enhancer Elements, Genetic; D005091:Exons; D005814:Genes, Viral; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D008969:Molecular Sequence Data; D017354:Point Mutation; D012326:RNA Splicing / Q000502:physiology*; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012261:Ribonucleoproteins / Q000502:physiology*; D014764:Viral Proteins / Q000502:physiology*
D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D012367:RNA, Viral; D012261:Ribonucleoproteins; D014764:Viral Proteins