The transcriptional cycle of HIV-1 in real-time and live cells

Boireau, Stéphanie; Maiuri, Paolo; Basyuk, Eugénia; Mata, Manuel de la; Knezevich, Anna; Pradet-Balade, Bérangère; Baecker, Volker; Kornblihtt, Alberto R.; Marcello, Alessandro; Bertrand, Édouard

doi:10.1083/jcb.200706018

Cited by 177 publications

(216 citation statements)

References 53 publications

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“…First, we chose RPB1 (the largest subunit of Pol II) known to have stable interactions with chromatin (Kimura et al , 2002; Hieda et al , 2005; Boireau et al , 2007; Darzacq et al , 2007; Fromaget & Cook, 2007) and also highly dynamic nuclear factors, like the TFIID subunit TAF5 and the GTF TFIIB (Chen et al , 2002; Sprouse et al , 2008; de Graaf et al , 2010; Ihalainen et al , 2012). In addition, eGFP was used as a control for a freely diffusing protein that should exhibit no specific interactions with the nuclear environment.…”

Section: Resultsmentioning

confidence: 99%

Coactivators and general transcription factors have two distinct dynamic populations dependent on transcription

et al. 2017

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SAGA and ATAC are two distinct chromatin modifying co‐activator complexes with distinct enzymatic activities involved in RNA polymerase II (Pol II) transcription regulation. To investigate the mobility of co‐activator complexes and general transcription factors in live‐cell nuclei, we performed imaging experiments based on photobleaching. SAGA and ATAC, but also two general transcription factors (TFIID and TFIIB), were highly dynamic, exhibiting mainly transient associations with chromatin, contrary to Pol II, which formed more stable chromatin interactions. Fluorescence correlation spectroscopy analyses revealed that the mobile pool of the two co‐activators, as well as that of TFIID and TFIIB, can be subdivided into “fast” (free) and “slow” (chromatin‐interacting) populations. Inhibiting transcription elongation decreased H3K4 trimethylation and reduced the “slow” population of SAGA, ATAC, TFIIB and TFIID. In addition, inhibiting histone H3K4 trimethylation also reduced the “slow” populations of SAGA and ATAC. Thus, our results demonstrate that in the nuclei of live cells the equilibrium between fast and slow population of SAGA or ATAC complexes is regulated by active transcription via changes in the abundance of H3K4me3 on chromatin.

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Section: Resultsmentioning

confidence: 99%

Coactivators and general transcription factors have two distinct dynamic populations dependent on transcription

et al. 2017

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“…Plasmids and Antibodies-The pTRIP-MS2ϫ24 and pMS2-YFP plasmids were described previously (47,48). TiVamp-CFP, Rab7-YFP, Gag-CFP, and Gag-TC were gifts of T. Galli (52), M. Zerial (49), H. G. Krausslich (50), and D. E. Ott (51).…”

Section: Methodsmentioning

confidence: 99%

Endosomal Trafficking of HIV-1 Gag and Genomic RNAs Regulates Viral Egress

Molle

Segura-Morales

Camus

et al. 2009

Journal of Biological Chemistry

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HIV-1 Gag can assemble and generate virions at the plasma membrane, but it is also present in endosomes where its role remains incompletely characterized. Here, we show that HIV-1 RNAs and Gag are transported on endosomal vesicles positive for TiVamp, a v-SNARE involved in fusion events with the plasma membrane. Inhibition of endosomal traffic did not prevent viral release. However, inhibiting lysosomal degradation induced an accumulation of Gag in endosomes and increased viral production 7-fold, indicating that transport of Gag to lysosomes negatively regulates budding. This also suggested that endosomal Gag-RNA complexes could access retrograde pathways to the cell surface and indeed, depleting cells of TiVampreduced viral production. Moreover, inhibition of endosomal transport prevented the accumulation of Gag at sites of cellular contact. HIV-1 Gag could thus generate virions using two pathways, either directly from the plasma membrane or through an endosome-dependent route. Endosomal Gag-RNA complexes may be delivered at specific sites to facilitate cell-to-cell viral transmission.

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“…Because an essential part of the TGS-AS mechanism is inhibition of RNAPII elongation, we first evaluated how SYNE2 E107 responds to activators and inhibitors of elongation. The chromatin-relaxing drugs trichostatin A (TSA) and 5 aza deoxycytidine (5azadC) are known to affect alternative splicing by promoting elongation (3, 9, 24), whereas the topoisomerase inhibitor camptothecin (CPT) was shown to inhibit elongation (24)(25)(26)(27). Unlike the exon E33 of fibronectin gene (FN1), where reduction in elongation promotes inclusion (Fig.…”

Section: Ago1 Preferentially Associates With Active Enhancers But Doementioning

confidence: 99%

Argonaute-1 binds transcriptional enhancers and controls constitutive and alternative splicing in human cells

Alló

Agirre

Bessonov

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

103

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The roles of Argonaute proteins in cytoplasmic microRNA and RNAi pathways are well established. However, their implication in small RNA-mediated transcriptional gene silencing in the mammalian cell nucleus is less understood. We have recently shown that intronic siRNAs cause chromatin modifications that inhibit RNA polymerase II elongation and modulate alternative splicing in an Argonaute-1 (AGO1)-dependent manner. Here we used chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) to investigate the genome-wide distribution of AGO1 nuclear targets. Unexpectedly, we found that about 80% of AGO1 clusters are associated with cell-type-specific transcriptional enhancers, most of them (73%) overlapping active enhancers. This association seems to be mediated by long, rather than short, enhancer RNAs and to be more prominent in intragenic, rather than intergenic, enhancers. Paradoxically, crossing ChIP-seq with RNA-seq data upon AGO1 depletion revealed that enhancer-bound AGO1 is not linked to the global regulation of gene transcription but to the control of constitutive and alternative splicing, which was confirmed by an individual gene analysis explaining how AGO1 controls inclusion levels of the cassette exon 107 in the SYNE2 gene.Argonaute proteins | transcriptional enhancers | alternative splicing A lternative splicing was initially seen as an interesting mechanism to explain protein diversity but affecting a limited number of mammalian genes. The recent development of high-throughput sequencing technologies has dramatically changed this view, generating a renewed interest in alternative splicing. We now know that alternative splicing affects transcripts from more than 90% of human genes (1) and that normal and pathological cell differentiation not only depends on differential gene expression but also on alternative splicing patterns. Mutations in alternative splicing regulatory sequences and factors are involved in the etiology of numerous hereditary diseases, premature aging, and cancer (2).Recently, amid an avalanche of papers reporting various connections between the chromatin context and splicing (3-9), a relationship between splicing and small RNAs has emerged. The convergence of these previously unrelated areas (RNA interference, chromatin, and splicing) has been studied by our laboratory, showing that siRNAs (20-25 nt long) targeting both intronic and exonic regions near the cassette exon 33 (E33, also known as EDI) of the fibronectin gene were able to regulate its alternative splicing by affecting the chromatin context at the target region, with an increase of histone tail modifications associated with gene silencing (H3K9me2 and H3K27me3, i.e., dimethylation of lysine 9 and trimethylation of lysine 27 of histone H3 respectively). Moreover, this effect was shown to be dependent on Argonaute proteins (AGO1 and AGO2) and involves a decrease of RNA polymerase II (RNAPII) elongation, which concomitantly up-regulates E33 inclusion into the mature mRNA (3). More recently, a similar effect was fou...

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The transcriptional cycle of HIV-1 in real-time and live cells

Cited by 177 publications

References 53 publications

Coactivators and general transcription factors have two distinct dynamic populations dependent on transcription

Coactivators and general transcription factors have two distinct dynamic populations dependent on transcription

Endosomal Trafficking of HIV-1 Gag and Genomic RNAs Regulates Viral Egress

Argonaute-1 binds transcriptional enhancers and controls constitutive and alternative splicing in human cells

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