Regulation of HIV-1 splicing

Human immunodeficiency virus type-1 (HIV-1) produces a single primary RNA transcript. The full-length transcript functions as RNA genome that is packaged into virions and as mRNA for translation of the Gag and Pol proteins. HIV-1 RNA contains several splice donor (5’splice site; 5’ss) and splice acceptor (3’splice site) sites. Differential usage of these sites results in the production of a variety of spliced RNAs that encode the other viral proteins. Splicing has to be strictly regulated for the balanced production of all viral RNAs and proteins. In this thesis, splicing control at the major 5’ss that is used for the synthesis of all spliced RNAs is investigated. We demonstrate that the efficiency of splicing at this 5’ss is modulated by the local RNA structure (i.e. the splice-donor [SD] RNA hairpin) and by the binding of regulatory SR proteins. Moreover, excessive splicing is prevented by the suboptimal sequence complementarity between the 5’ss and the cellular U1 snRNA, which limits binding of this essential splice factor. All three regulatory features of the 5’ss region are highly conserved among different virus isolates, which supports their importance. Furthermore, we demonstrate that the viral Tat protein, which is known to activate transcription from the HIV-1 LTR promoter, also stimulates splicing at the major 5’ss. Our results indicate that HIV-1 transcription and splicing are tightly coupled processes through the coordinated action of Tat. We also set out to identify 5’ss regions in human RNA transcripts that are controlled by local RNA structure.