RU5 of Mason-Pfizer Monkey Virus 5′ Long Terminal Repeat Enhances Cytoplasmic Expression of Human Immunodeficiency Virus Type 1 gag-pol and Nonviral Reporter RNA

Abstract

Retroviruses utilize an unspliced version of their primary transcription product as an RNA template for synthesis of viral Gag and Pol structural and enzymatic proteins. Cytoplasmic expression of the gag-pol RNA is achieved despite the lack of intron removal and the presence of a long and highly structured 5 untranslated region that inhibits efficient ribosome scanning. In this study, we have identified for the first time that the 5 long terminal repeat ( Retroviruses need to subvert typical cellular posttranscriptional control mechanisms to produce progeny virions. Typical pre-mRNAs are assembled in ribonucleoprotein (RNP) complexes during transcription and RNA maturation that qualify processed transcripts for nuclear export and efficient cytoplasmic expression (11,24,26,30). By contrast, retroviral premRNA recruits viral or cellular posttranscriptional modulators that activate efficient nuclear export and cytoplasmic expression despite lack of intron removal (2,12,25,44). Once in the cytoplasm, the unspliced viral transcript exhibits dual function as mRNA template for translation of Gag precursor protein and Gag-Pol polyprotein and as genomic RNA that is packaged into progeny virions (6). The RNA packaging signal is a series of RNA structural motifs within the 5Ј untranslated region (UTR) that are recognized by the Gag nucleocapsid protein, which directs assembly of the unspliced RNA into progeny virions (1, 35). Structured 5Ј UTRs typically inhibit efficient translation by steric hindrance of ribosome scanning from the 5Ј cap to the proximal start codon (19,29,34). Mutational analysis of structural motifs in the human immunodeficiency virus type 1 (HIV-1) 5Ј UTR has verified that they inhibit efficient translation (18,31,33). Thus, the unspliced viral RNA subverts typical barriers to both nuclear export and efficient translation to achieve productive cytoplasmic expression of viral structural and enzymatic proteins.HIV-1 is a complex retrovirus that encodes the viral posttranscriptional regulatory protein Rev. Rev interacts with newly synthesized viral RNA (22) at the Rev-responsive element (RRE) present in distal intronic sequences to activate nuclear export despite lack of intron removal (8,20,21,28,46). Rev connects RRE-containing RNA to the CRM1/exportin 1 nuclear export receptor, which is typically utilized by 5S rRNA and shuttling proteins that contain a leucine-rich nuclear export sequence (17, 32). Genetically simpler retroviruses lack a viral Rev protein. However, the type D Mason-Pfizer monkey virus (MPMV) and simian retrovirus type 1 contain a functional homolog of RRE, which is designated the constitutive transport element (CTE). The CTE is a redundant stem-loop structure that is positioned in the 3Ј UTR and facilitates nuclear export of intron-containing RNA in transfected cells (5,15,16,38,42). CTE facilitates nuclear export by interaction with Tap and cofactor NXT1, which have been implicated as global modulators of the nuclear export of fully processed cellular mRNAs (4,25,44). Recently, studi...

Jaagsiekte Sheep Retrovirus Encodes a Regulatory Factor, Rej, Required for Synthesis of Gag Protein

“…Further, the Tap protein could be immunoprecipitated from the polyribosome fractions of sucrose sedimentation gradients, indicating that Tap associates with polyribosomes (29). Similarly, cellular RNA helicase A (RHA) has recently been shown to enhance translation of retroviral RNAs containing a 5Ј-terminal posttranscriptional control element, found in avian spleen necrosis virus (5) and MPMV (25). RHA binds the highly structured posttranscriptional control element to mediate the translational enhancement (21).…”

A novel role of RNA helicase A in regulation of translation of type I collagen mRNAs

“…PCE is a highly structured, orientationdependent motif present in the 59 UTRs of some retroviral mRNAs (Butsch et al 1999;Roberts and Boris-Lawrie 2000;Hull and Boris-Lawrie 2002). The 59PCE activity is positively regulated by RHA and is essential for the translation efficiency of viral mRNAs (Butsch et al 1999;Hull and Boris-Lawrie 2002). In addition, Hartman et al (2006) demonstrated that the PCE-like element in the 59 UTR of human JunD mRNA interacts with RHA and regulates translation.…”

“…Recently, it was demonstrated that RHA is a necessary cofactor to promote translation initiation of highly structured mRNAs containing the post-transcriptional control element (PCE) Zhang and Grosse 2004;Hartman et al 2006). PCE is a highly structured, orientationdependent motif present in the 59 UTRs of some retroviral mRNAs (Butsch et al 1999;Roberts and Boris-Lawrie 2000;Hull and Boris-Lawrie 2002). The 59PCE activity is positively regulated by RHA and is essential for the translation efficiency of viral mRNAs (Butsch et al 1999;Hull and Boris-Lawrie 2002).…”

Primary Sequence and Secondary Structure Motifs in Spleen Necrosis Virus RU5 Confer Translational Utilization of Unspliced Human Immunodeficiency Virus Type 1 Reporter RNA

“…The 3Ј UTRs of avian leukosis virus and the related Rous sarcoma virus contain one and two copies, respectively, of a direct repeat (DR) element that is necessary for cytoplasmic accumulation and stability of unspliced viral RNA and for assembly of progeny virions (1, 22-24, 29, 30, 33). The 5Ј long terminal repeats (LTRs) of spleen necrosis virus (SNV) and MPMV contain a unique posttranscriptional control element that facilitates reporter gene expression from unspliced human immunodeficiency virus type 1 (HIV-1) gag reporter RNA independently of Rev/Rev-responsive element (RRE), and also nonviral luc RNA (6,15,28).Extensive mutagenesis studies of CTE and DR have mapped necessary and redundant structural motifs that present unpaired nucleotides for interaction with cellular posttranscriptional modulators. The MPMV and SRV-1 CTEs share 92% sequence homology and are 154-nucleotide (nt) orientationand position-dependent RNA elements (27, 32).…”