The D1-A2 and D2-A2 pairs of splice sites from human immunodeficiency virus type 1 are highly efficient in vitro, in spite of an unusual branch site.

Using in vitro splicing assays with HeLa cell nuclear extracts, we showed that the HIV-1 pairs of splice sites D1-A2 and D2-A2 are efficiently used in vitro, as compared to the control D1-A2 pair of sites from the E3 transcription unit of human adenovirus-2. The strong efficiency of the two HIV-1 pairs of sites is surprising, as we also showed by primer extension analysis that the branch-site sequence used at the HIV-1 acceptor site A2 is UAGCAGA, with a dominant utilization of the ultimate G as the branched residue. No significant increase of the splicing efficiency was observed upon replacement of the wild-type branch-site sequence by a canonical sequence, in spite of the utilization of an A residue as the branched nucleotide. Results are discussed taking into account the present knowledge on branch-site selection.