UC San Diego

Alternative splicing is critical to many cellular functions and appears to be primarily regulated through co -transcriptional mechanisms, where spliced-transcript identity is determined during transcriptional elongation. For HIV-1, alternative splicing is essential since the viral genome encodes multiple overlapping reading frames. Using quantitative single-cell imaging, we find that HIV-1 splicing is not co-transcriptionally regulated. Strikingly, HIV-1 mRNA appears to be spliced post-transcriptionally in a serial cascade. We propose a sensitive new assay to detect in vivo post-transcriptional splicing. The assay result shows that HIV-1 mRNA does not require transcription for spliceosome assembly. This serial splicing cascade appears crucial for the HIV-1 Rev negative-feedback circuit and in agreement with mathematical models. We experimentally verify the counter- intuitive prediction that Rev over-expression leads to reduced HIV-1 p24 expression. Serial post-transcriptional splicing may provide a novel target for antivirals and, more generally, an alternate gene-regulatory mechanism