Small-Molecule Inhibition of HIV pre-mRNA Splicing as a Novel Antiretroviral Therapy to Overcome Drug Resistance

Bakkour, Nadia; Lin, Yea-Lih; Maire, Sophie; Ayadi, Lilia; Mahuteau-Betzer, Florence; Nguyen, Chi Hung; Mettling, Clément; Portalès, Pierre; Grierson, David S.; Chabot, Benoît; Jeanteur, Philippe; Branlant, Christiane; Corbeau, Pierre; Tazi, Jamal

doi:10.1371/journal.ppat.0030159

Cited by 78 publications

(110 citation statements)

References 42 publications

Supporting

Mentioning

104

Contrasting

Unclassified

Order By: Relevance

“…gRNA regulation diverges from that of most cellular mRNAs, which may present novel opportunities for antiretroviral drug development. Small molecule inhibitors have been developed for SR proteins, and these have been proposed to be antiretroviral compounds, though mostly as splicing inhibitors (2,40,70). Identifying the mechanism by which SRp40/55 regulate HIV-1 Gag translation may lead to the development of specific small molecule inhibitors that inhibit HIV-1 protein production.…”

Section: Discussionmentioning

confidence: 99%

SRp40 and SRp55 Promote the Translation of Unspliced Human Immunodeficiency Virus Type 1 RNA

Swanson

Sherer

Malim

2010

J Virol

View full text Add to dashboard Cite

Nuclear RNA processing events, such as 5 cap formation, 3 polyadenylation, and pre-mRNA splicing, mark mRNA for efficient translation. Splicing enhances translation via the deposition of the exon-junction complex and other multifunctional splicing factors, including SR proteins. All retroviruses synthesize their structural and enzymatic proteins from unspliced genomic RNAs (gRNAs) and must therefore exploit unconventional strategies to ensure their effective expression. Here, we report that specific SR proteins, particularly SRp40 and SRp55, promote human immunodeficiency virus type 1 (HIV-1) Gag translation from unspliced (introncontaining) viral RNA. This activity does not correlate with nucleocytoplasmic shuttling capacity and, in the case of SRp40, is dependent on the second RNA recognition motif and the arginine-serine (RS) domain. While SR proteins enhance Gag expression independent of RNA nuclear export pathway choice, altering the nucleotide sequence of the gag-pol coding region by codon optimization abolishes this effect. We therefore propose that SR proteins couple HIV-1 gRNA biogenesis to translational utilization.From transcription to translation to cytoplasmic mRNA degradation, the sequential phases of gene expression are coupled physically and functionally. In humans, 94% of genes have more than one exon (79) and pre-mRNA splicing plays a central role in regulating transcription, nuclear stability, 3Ј end formation, nuclear export, cytoplasmic trafficking, cytoplasmic stability, translation efficiency, and even posttranslational events, such as protein half-life (reviewed in references 56 and 75). The integration of pre-mRNA splicing with later steps in RNA metabolism can prevent the translation of deleterious RNAs in at least two ways. First, mRNAs containing functional introns are retained by the nucleus (14, 45), thereby preventing the translation of incompletely processed transcripts and the synthesis of deleterious protein products. Second, and less well understood, the coupling of splicing to the control of nuclear stability, export, and translation prevents the translation of unspliced parasitic and/or noncoding RNAs (reviewed in reference 6). Because viruses frequently depend upon the translation of unspliced viral RNAs, they need to have developed mechanisms to circumvent such barriers to gene expression.All retroviruses use a combination of spliced and unspliced RNAs to express their proteins (reviewed in references 20 and 71). The nuclear export of a full-length, unspliced, introncontaining transcript is essential for the replication of all retroviruses, since this transcript is (i) the viral genome (genomic RNA [gRNA]) that is packaged into virions, (ii) the mRNA that is translated into Gag and Gag-polymerase (Pol), the polyproteins that yield the virion structural and enzymatic proteins, and (iii) a physical scaffold that helps promote virion assembly. Human immunodeficiency virus type 1 (HIV-1) has one of the most complicated gene expression strategies of all known retroviruses, where...

show abstract

Section: Discussionmentioning

confidence: 99%

SRp40 and SRp55 Promote the Translation of Unspliced Human Immunodeficiency Virus Type 1 RNA

Swanson

Sherer

Malim

2010

J Virol

View full text Add to dashboard Cite

show abstract

“…The dependence of viruses on ASF/SF2 and its oncogenic potential makes ASF/SF2 an attractive target for therapy in infections and cancer. Indeed, IDC-16, a small molecule inhibitor of ASF/SF2 activity, was shown to reduce human immunodeficiency virus replication (40). Furthermore, ASF/SF2 was specifically up-regulated in a cell line model of cervical tumor progression (41), suggesting its potential as a biomarker for cancer.…”

Section: Discussionmentioning

confidence: 99%

Alternative Splicing Factor/Splicing Factor 2 Regulates the Expression of the ζ Subunit of the Human T Cell Receptor-associated CD3 Complex

Moulton

Tsokos

2010

Journal of Biological Chemistry

View full text Add to dashboard Cite

T cells from patients with systemic lupus erythematosus express decreased levels of the T cell receptor-associated CD3 chain, a feature directly linked to their aberrant function. The decrease in CD3 protein expression is in part due to decreased levels of functional wild type isoform of the 3-untranslated region (UTR) of CD3 mRNA with concomitant increased levels of an unstable alternatively spliced isoform. In order to identify factors involved in the post-transcriptional regulation of CD3, we performed mass spectrometric analysis of Jurkat T cell nuclear proteins "pulled down" by a CD3 3-UTR oligonucleotide, which identified the splicing protein alternative splicing factor/splicing factor 2 (ASF/SF2). We show for the first time that ASF/SF2 binds specifically to the 3-UTR of CD3 and regulates expression of CD3 protein by limiting the production of the alternatively spliced isoform. During activation of human T cells, an increase in the wild type CD3 mRNA is associated with increased expression of ASF/SF2. Finally, we show a significant correlation between ASF/SF2 and CD3 protein levels in T cells from systemic lupus erythematosus patients. Thus, our results identify ASF/SF2 as a novel factor in the regulation of alternative splicing of the 3-UTR of CD3 and protein expression in human T cells. Systemic lupus erythematosus (SLE)2 is a chronic multisystem autoimmune disease characterized by abnormal cellular and humoral immune responses and a disease course marked by relapses (flares) and remissions. A key signaling defect of T cells in SLE patients is their aberrant expression of the T cell receptor (TCR)-associated CD3 chain, the crucial signaling transducer of the TCR (1). CD3 protein expression is heterogeneous in SLE patients and tends to inversely correlate with severity of the disease. Consequently, many patients exhibit decreased (20 -80% of normal) levels of CD3 protein, whereas others express normal levels of CD3 protein when compared with healthy individuals. The homologous Fc receptor ␥ chain replaces CD3 chain in the TCR CD3 complex in these CD3-low T cells (2) and recruits the spleen tyrosine kinase (Syk) instead of the normally recruited -associated protein (ZAP-70) (3). This "rewiring" of the TCR, along with preclustered TCRbearing lipid rafts, leads to aberrantly increased phosphorylation and calcium flux upon activation (4). Despite this overexcitable phenotype, SLE T cells fail to produce the vital cytokine interleukin-2. Replenishment of the CD3 in the SLE T cells in vitro reverts this phenotype and more importantly restores interleukin-2 production (5). Although these findings highlight a central role for the aberrant CD3 expression in the SLE T cell defect, the mechanisms regulating the expression of CD3 chain in physiology and in disease are not fully understood.The CD3 gene spans 31 kb in the chromosome 1q23.1 locus, a region associated with SLE susceptibility (6), and bears eight exons separated by introns ranging from 700 bp to greater than 8 kb (Fig. 1A, top) (7,8). The CD3 mRNA is a 1...

show abstract

“…For example, indole derivative compounds (IDCs) have been identified that selectively inhibit SR protein family members (Soret et al, 2005). Certain IDCs potently inhibited HIV-1 replication in cell culture experiments (Bakkour et al, 2007;Soret et al, 2005). Other IDCs inhibit replication of murine leukemia virus (MLV) and protect mice from MLV-induced pathogenesis (Keriel et al, 2009).…”

Section: Srp40mentioning

confidence: 99%

HIV-1 splicing is controlled by local RNA structure and binding of splicing regulatory proteins at the major 5′ splice site

Müeller

Berkhout

Das

2015

Journal of General Virology

View full text Add to dashboard Cite

The 59 leader region of the human immunodeficiency virus 1 (HIV-1) RNA genome contains the major 59 splice site (ss) that is used in the production of the many spliced viral RNAs. This splice-donor (SD) region can fold into a stable stem-loop structure and the thermodynamic stability of this RNA hairpin influences splicing efficiency. In addition, splicing may be modulated by binding of splicing regulatory (SR) proteins, in particular SF2/ASF (SRSF1), SC35 (SRSF2), SRp40 (SRSF5) and SRp55 (SRSF6), to sequence elements in the SD region. The role of RNA structure and SR protein binding in splicing control was previously studied by functional analysis of mutant SD sequences. The interpretation of these studies was complicated by the fact that most mutations simultaneously affect both structure and sequence elements. We therefore tried to disentangle the contribution of these two variables by designing more precise SD region mutants with a single effect on either the sequence or the structure. The current analysis indicates that HIV-1 splicing at the major 59ss is modulated by both the stability of the local RNA structure and the binding of splicing regulatory proteins.

show abstract

Small-Molecule Inhibition of HIV pre-mRNA Splicing as a Novel Antiretroviral Therapy to Overcome Drug Resistance

Cited by 78 publications

References 42 publications

SRp40 and SRp55 Promote the Translation of Unspliced Human Immunodeficiency Virus Type 1 RNA

SRp40 and SRp55 Promote the Translation of Unspliced Human Immunodeficiency Virus Type 1 RNA

Alternative Splicing Factor/Splicing Factor 2 Regulates the Expression of the ζ Subunit of the Human T Cell Receptor-associated CD3 Complex

HIV-1 splicing is controlled by local RNA structure and binding of splicing regulatory proteins at the major 5′ splice site

Contact Info

Product

Resources

About