Activation of HIV-1 pre-mRNA 3′ processing in vitro requires both an upstream element and TAR.

Abstract

The architecture of the human immunodeficiency virus type 1 (HIV‐1) genome presents an intriguing dilemma for the 3′ processing of viral transcripts‐‐to disregard a canonical ‘core’ poly(A) site processing signal present at the 5′ end of the transcript and yet to utilize efficiently an identical signal that resides at the 3′ end of the message. The choice of processing sites in HIV‐1 appears to be influenced by two factors: (i) proximity to the cap site, and (ii) sequences upstream of the core poly(A) site. We now demonstrate that an in vivo‐defined upstream element that resides within the U3 region, 76 nucleotides upstream of the AAUAAA hexamer, acts specifically to enhance 3′ processing at the HIV‐1 core poly(A) site in vitro. We furthermore show that efficient in vitro 3′ processing requires the RNA stem‐loop structure of TAR, which serves to juxtapose spatially the upstream element and the core poly(A) site. An analysis of the stability of 3′ processing complexes formed at the HIV‐1 poly(A) site in vitro suggests that the upstream element may function by increasing processing complex stability at the core poly(A) site.

3′-end-forming signals of yeast mRNA

“…In addition to the AAUAAA motif and the downstream GU-rich or U-rich element of mammalian systems, upstream sequences that contribute to the efficiency of mRNA 3Ј end formation were also identified in the mammalian viruses simian virus 40 (6,36), adenovirus (8,9), hepatitis B virus (29), and human immunodeficiency virus type 1 (3,11,12,38,39). These elements function in an orientation-and position-dependent manner, and several appear to be functionally analogous.…”

Protein factors in pre-mRNA 3′-end processing

“…Mutations of the PAS abolished the cross-linking of nuclear extract [89], while they only have a small effect on the cross-linking of the recombinant CPSF-160 [88], suggesting that other proteins are required for the highly specific binding to the PAS by CPSF-160. CPSF-160 can also bind at a site other than the PAS in HIV pre-mRNA [90].…”

HIV-1 as RNA evolution machine

“…This fine-tuning of the polyA hairpin stability sets the stage for the regulation of HIV-1 RNA polyadenylation, which involves additional suppressive mechanisms for the 5' copy and activation mechanisms for the 3' copy. [54][55][56][57][58][59][60][61] Regulation of HIV-1 RNA Splicing…”