Imagerie

@article{pmid37254203,

title = {DNA topoisomerase 1 represses HIV-1 promoter activity through its interaction with a guanine quadruplex present in the LTR sequence},

author = {María José Lista and Anne-Caroline Jousset and Mingpan Cheng and Violaine Saint-André and Elouan Perrot and Melissa Rodrigues and Carmelo Di Primo and Danielle Gadelle and Elenia Toccafondi and Emmanuel Segeral and Clarisse Berlioz-Torrent and Stéphane Emiliani and Jean-Louis Mergny and Marc Lavigne},

doi = {10.1186/s12977-023-00625-8},

issn = {1742-4690},

year  = {2023},

date = {2023-05-01},

urldate = {2023-05-01},

journal = {Retrovirology},

volume = {20},

number = {1},

pages = {10},

abstract = {BACKGROUND: Once integrated in the genome of infected cells, HIV-1 provirus is transcribed by the cellular transcription machinery. This process is regulated by both viral and cellular factors, which are necessary for an efficient viral replication as well as for the setting up of viral latency, leading to a repressed transcription of the integrated provirus.nnRESULTS: In this study, we examined the role of two parameters in HIV-1 LTR promoter activity. We identified DNA topoisomerase1 (TOP1) to be a potent repressor of this promoter and linked this repression to its catalytic domain. Additionally, we confirmed the folding of a Guanine quadruplex (G4) structure in the HIV-1 promoter and its repressive effect. We demonstrated a direct interaction between TOP1 and this G4 structure, providing evidence of a functional relationship between the two repressive elements. Mutations abolishing G4 folding affected TOP1/G4 interaction and hindered G4-dependent inhibition of TOP1 catalytic activity in vitro. As a result, HIV-1 promoter activity was reactivated in a native chromatin environment. Lastly, we noticed an enrichment of predicted G4 sequences in the promoter of TOP1-repressed cellular genes.nnCONCLUSIONS: Our results demonstrate the formation of a TOP1/G4 complex on the HIV-1 LTR promoter and its repressive effect on the promoter activity. They reveal the existence of a new mechanism of TOP1/G4-dependent transcriptional repression conserved between viral and human genes. This mechanism contrasts with the known property of TOP1 as global transcriptional activator and offers new perspectives for anti-cancer and anti-viral strategies.},

keywords = {MARQUET, NEGRONI, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{pmid36161446,

title = {The DEAD box RNA helicase DDX42 is an intrinsic inhibitor of positive-strand RNA viruses},

author = {Boris Bonaventure and Antoine Rebendenne and Ana Luiza Chaves Valadão and Mary Arnaud-Arnould and Ségolène Gracias and Francisco Garcia de Gracia and Joe McKellar and Emmanuel Labaronne and Marine Tauziet and Valérie Vivet-Boudou and Eric Bernard and Laurence Briant and Nathalie Gros and Wassila Djilli and Valérie Courgnaud and Hugues Parrinello and Stéphanie Rialle and Mickaël Blaise and Laurent Lacroix and Marc Lavigne and Jean-Christophe Paillart and Emiliano P Ricci and Reiner Schulz and Nolwenn Jouvenet and Olivier Moncorgé and Caroline Goujon},

url = {https://pubmed.ncbi.nlm.nih.gov/36161446/},

doi = {10.15252/embr.202154061},

issn = {1469-3178},

year  = {2022},

date = {2022-09-01},

urldate = {2022-09-01},

journal = {EMBO Rep},

pages = {e54061},

abstract = {Genome-wide screens are powerful approaches to unravel regulators of viral infections. Here, a CRISPR screen identifies the RNA helicase DDX42 as an intrinsic antiviral inhibitor of HIV-1. Depletion of endogenous DDX42 increases HIV-1 DNA accumulation and infection in cell lines and primary cells. DDX42 overexpression inhibits HIV-1 infection, whereas expression of a dominant-negative mutant increases infection. Importantly, DDX42 also restricts LINE-1 retrotransposition and infection with other retroviruses and positive-strand RNA viruses, including CHIKV and SARS-CoV-2. However, DDX42 does not impact the replication of several negative-strand RNA viruses, arguing against an unspecific effect on target cells, which is confirmed by RNA-seq analysis. Proximity ligation assays show DDX42 in the vicinity of viral elements, and cross-linking RNA immunoprecipitation confirms a specific interaction of DDX42 with RNAs from sensitive viruses. Moreover, recombinant DDX42 inhibits HIV-1 reverse transcription in vitro. Together, our data strongly suggest a direct mode of action of DDX42 on viral ribonucleoprotein complexes. Our results identify DDX42 as an intrinsic viral inhibitor, opening new perspectives to target the life cycle of numerous RNA viruses.},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{Libre2022,

title = {A Conserved uORF Regulates APOBEC3G Translation and Is Targeted by HIV-1 Vif Protein to Repress the Antiviral Factor},

author = {C Libre and T Seissler and S Guerrero and J Batisse and C Verriez and B Stupfler and O Gilmer and R Cabrera-Rodriguez and M Weber and A Valenzuela-Fernandez and A Cimarelli and L Etienne and R Marquet and J C Paillart},

url = {https://www.mdpi.com/2227-9059/10/1/13},

doi = {10.1101/2021.01.13.426487},

year  = {2022},

date = {2022-01-01},

urldate = {2022-01-01},

journal = {Biomedicines},

volume = {10},

number = {1},

pages = {13},

abstract = {The HIV-1 Vif protein is essential for viral fitness and pathogenicity. Vif decreases expression of cellular cytosine deaminases APOBEC3G (A3G), A3F, A3D and A3H, which inhibit HIV-1 replication by inducing hypermutations during reverse transcription. Vif counteracts A3G by several non-redundant mechanisms (transcription, translation and protein degradation) that concur in reducing the levels of A3G in cell and in preventing its incorporation into viral particles. How Vif affects A3G translation remains unclear. Here, we uncovered the importance of a short conserved uORF (upstream ORF) located within two critical stem-loop structures of the 5-untranslated region (5-UTR) of A3G mRNA. Extensive mutagenesis of A3G 5-UTR, combined with an analysis of their translational effect in transfected cells, indicated that the uORF represses A3G translation and that A3G mRNA is translated through a dual leaky-scanning and re-initiation mechanism. Interestingly, the uORF is also mandatory for the Vif-mediated repression of A3G translation. Furthermore, we showed that the redirection of A3G mRNA into stress granules was dependent not only on Vif, but also on the uORF. Overall, we discovered that A3G translation is regulated by a small uORF conserved in the human population and that Vif uses this specific motif to repress its translation.},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{nokey,

title = {Structural maturation of the HIV-1 RNA 5' untranslated region by Pr55(Gag) and its maturation products},

author = {O. Gilmer and E. Mailler and J. C. Paillart and A. Mouhand and C. Tisne and J. Mak and R. P. Smyth and R. Marquet and V. Vivet-Boudou},

url = {https://www.tandfonline.com/doi/full/10.1080/15476286.2021.2021677},

isbn = {35067194},

year  = {2022},

date = {2022-01-01},

journal = {RNA Biol},

volume = {19},

number = {1},

pages = {191-205},

abstract = {Maturation of the HIV-1 viral particles shortly after budding is required for infectivity. During this process, the Pr55(Gag) precursor undergoes a cascade of proteolytic cleavages, and whilst the structural rearrangements of the viral proteins are well understood, the concomitant maturation of the genomic RNA (gRNA) structure is unexplored, despite evidence that it is required for infectivity. To get insight into this process, we systematically analysed the interactions between Pr55(Gag) or its maturation products (NCp15, NCp9 and NCp7) and the 5' gRNA region and their structural consequences, in vitro. We show that Pr55(Gag) and its maturation products mostly bind at different RNA sites and with different contributions of their two zinc knuckle domains. Importantly, these proteins have different transient and permanent effects on the RNA structure, the late NCp9 and NCp7 inducing dramatic structural rearrangements. Altogether, our results reveal the distinct contributions of the different Pr55(Gag) maturation products on the gRNA structural maturation.},

note = {1555-8584 (Electronic) 

1547-6286 (Linking) 

Journal Article},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{nokey,

title = {Visualization of Retroviral Gag-Genomic RNA Cellular Interactions Leading to Genome Encapsidation and Viral Assembly: An Overview},

author = {S. Bernacchi},

url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35215917},

doi = {10.3390/v14020324},

isbn = {35215917},

year  = {2022},

date = {2022-01-01},

journal = {Viruses},

volume = {14},

number = {2},

abstract = {Retroviruses must selectively recognize their unspliced RNA genome (gRNA) among abundant cellular and spliced viral RNAs to assemble into newly formed viral particles. Retroviral gRNA packaging is governed by Gag precursors that also orchestrate all the aspects of viral assembly. Retroviral life cycles, and especially the HIV-1 one, have been previously extensively analyzed by several methods, most of them based on molecular biology and biochemistry approaches. Despite these efforts, the spatio-temporal mechanisms leading to gRNA packaging and viral assembly are only partially understood. Nevertheless, in these last decades, progress in novel bioimaging microscopic approaches (as FFS, FRAP, TIRF, and wide-field microscopy) have allowed for the tracking of retroviral Gag and gRNA in living cells, thus providing important insights at high spatial and temporal resolution of the events regulating the late phases of the retroviral life cycle. Here, the implementation of these recent bioimaging tools based on highly performing strategies to label fluorescent macromolecules is described. This report also summarizes recent gains in the current understanding of the mechanisms employed by retroviral Gag polyproteins to regulate molecular mechanisms enabling gRNA packaging and the formation of retroviral particles, highlighting variations and similarities among the different retroviruses.},

note = {1999-4915 (Electronic) 

1999-4915 (Linking) 

Journal Article 

Review 

Research Support, Non-U.S. Gov't},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{C2021b,

title = {Post-Translational Modifications of Retroviral HIV-1 Gag Precursors: An Overview of Their Biological Role},

author = {C Busienne and R Marquet and J C Paillart and S Bernacchi},

url = {https://www.mdpi.com/1422-0067/22/6/2871},

doi = {10.3390/ijms22062871},

year  = {2021},

date = {2021-01-01},

journal = {Int. J. Mol. Sci.},

volume = {22},

number = {6},

pages = {2871},

abstract = {Protein post-translational modifications (PTMs) play key roles in eukaryotes since they finely regulate numerous mechanisms used to diversify the protein functions and to modulate their signaling networks. Besides, these chemical modifications also take part in the viral hijacking of the host, and also contribute to the cellular response to viral infections. All domains of the human immunodeficiency virus type 1 (HIV-1) Gag precursor of 55-kDa (Pr55Gag), which is the central actor for viral RNA specific recruitment and genome packaging, are post-translationally modified. In this review, we summarize the current knowledge about HIV-1 Pr55Gag PTMs such as myristoylation, phosphorylation, ubiquitination, sumoylation, methylation, and ISGylation in order to figure out how these modifications affect the precursor functions and viral replication. Indeed, in HIV-1, PTMs regulate the precursor trafficking between cell compartments and its anchoring at the plasma membrane, where viral assembly occurs. Interestingly, PTMs also allow Pr55Gag to hijack the cell machinery to achieve viral budding as they drive recognition between viral proteins or cellular components such as the ESCRT machinery. Finally, we will describe and compare PTMs of several other retroviral Gag proteins to give a global overview of their role in the retroviral life cycle.},

keywords = {HIV-1, MARQUET, PAILLART, post-translational modifications, Pr55Gag precursor, retroviral Gag precursors, retroviral life cycle, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{Chameettachal2021,

title = {A purine loop and the primer binding site are critical for the selective encapsidation of mouse mammary tumor virus genomic RNA by Pr77Gag},

author = {A Chameettachal and V Vivet-Boudou and F N Pitchai and Pillai V N and L M Ali and A Krishnan and S Bernacchi and F Mustafa and R Marquet and T A Rizvi},

year  = {2021},

date = {2021-01-01},

urldate = {2021-01-01},

journal = {Nucleic Acids Res},

volume = {49},

number = {8},

pages = {4668-4688},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{Stupfler2021,

title = {Degradation-independent inhibition of APOBEC3G by HIV-1 Vif protein},

author = {B Stupfler and C Verriez and S Gallois-Montbrun and R Marquet and J C Paillart},

url = {https://www.mdpi.com/1999-4915/13/4/617},

doi = {10.3390/v13040617},

year  = {2021},

date = {2021-01-01},

journal = {Viruses},

volume = {13},

number = {4},

pages = {617},

abstract = {The ubiquitin–proteasome system plays an important role in the cell under normal physiological conditions but also during viral infections. Indeed, many auxiliary proteins from the (HIV-1) divert this system to its own advantage, notably to induce the degradation of cellular restriction factors. For instance, the HIV-1 viral infectivity factor (Vif) has been shown to specifically counteract several cellular deaminases belonging to the apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC3 or A3) family (A3A to A3H) by recruiting an E3-ubiquitin ligase complex and inducing their polyubiquitination and degradation through the proteasome. Although this pathway has been extensively characterized so far, Vif has also been shown to impede A3s through degradation-independent processes, but research on this matter remains limited. In this review, we describe our current knowledge regarding the degradation-independent inhibition of A3s, and A3G in particular, by the HIV-1 Vif protein, the molecular mechanisms involved, and highlight important properties of this small viral protein.},

keywords = {APOBEC3G, deamination, encapsidation, HIV, MARQUET, PAILLART, proteasome, RNP granules, Translation, ubiquitin, Unité ARN, vif},

pubstate = {published},

tppubtype = {article}

}

@article{Welker2021,

title = {Importance of Viral Late Domains in Budding and Release of Enveloped RNA Viruses},

author = {L Welker and J C Paillart and S Bernacchi},

url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34452424},

doi = {10.3390/v13081559},

isbn = {34452424},

year  = {2021},

date = {2021-01-01},

urldate = {2021-01-01},

journal = {Viruses},

volume = {13},

number = {8},

pages = {1559},

abstract = {Late assembly (L) domains are conserved sequences that are necessary for the late steps of viral replication, acting like cellular adaptors to engage the ESCRT membrane fission machinery that promote virion release. These short sequences, whose mutation or deletion produce the accumulation of immature virions at the plasma membrane, were firstly identified within retroviral Gag precursors, and in a further step, also in structural proteins of many other enveloped RNA viruses including arenaviruses, filoviruses, rhabdoviruses, reoviruses, and paramyxoviruses. Three classes of L domains have been identified thus far (PT/SAP, YPXnL/LXXLF, and PPxY), even if it has recently been suggested that other motifs could act as L domains. Here, we summarize the current state of knowledge of the different types of L domains and their cellular partners in the budding events of RNA viruses, with a particular focus on retroviruses.},

note = {1999-4915 (Electronic) 

1999-4915 (Linking) 

Journal Article 

Review},

keywords = {PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{nokey,

title = {Chemical and Enzymatic Probing of Viral RNAs: From Infancy to Maturity and Beyond},

author = {O Gilmer and E Quignon and A C Jousset and J C Paillart and R Marquet and V Vivet-Boudou},

url = {https://www.mdpi.com/1999-4915/13/10/1894},

doi = {v13101894},

year  = {2021},

date = {2021-01-01},

urldate = {2021-01-01},

journal = {Viruses},

volume = {13},

number = {10},

pages = {1894},

abstract = {RNA molecules are key players in a variety of biological events, and this is particularly true for viral RNAs. To better understand the replication of those pathogens and try to block them, special attention has been paid to the structure of their RNAs. Methods to probe RNA structures have been developed since the 1960s; even if they have evolved over the years, they are still in use today and provide useful information on the folding of RNA molecules, including viral RNAs. The aim of this review is to offer a historical perspective on the structural probing methods used to decipher RNA structures before the development of the selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) methodology and to show how they have influenced the current probing techniques. Actually, these technological breakthroughs, which involved advanced detection methods, were made possible thanks to the development of next-generation sequencing (NGS) but also to the previous works accumulated in the field of structural RNA biology. Finally, we will also discuss how high-throughput SHAPE (hSHAPE) paved the way for the development of sophisticated RNA structural techniques.},

keywords = {Capillary electrophoresis, chemical probe, enzymatic probe, high-throughput sequencing, MARQUET, mutational profiling, PAILLART, RNA, SHAPE, structure, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{nokey,

title = {The HIV-1 Nucleocapsid Regulates Its Own Condensation by Phase-Separated Activity-Enhancing Sequestration of the Viral Protease during Maturation},

author = {S. Lyonnais and S. K. Sadiq and C. Lorca-Oro and L. Dufau and S. Nieto-Marquez and T. Escriba and N. Gabrielli and X. Tan and M. Ouizougun-Oubari and J. Okoronkwo and M. Reboud-Ravaux and J. M. Gatell and R. Marquet and J. C. Paillart and A. Meyerhans and C. Tisne and R. J. Gorelick and G. Mirambeau},

url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34835118},

doi = {10.3390/v13112312},

isbn = {34835118},

year  = {2021},

date = {2021-01-01},

journal = {Viruses},

volume = {13},

number = {11},

abstract = {A growing number of studies indicate that mRNAs and long ncRNAs can affect protein populations by assembling dynamic ribonucleoprotein (RNP) granules. These phase-separated molecular 'sponges', stabilized by quinary (transient and weak) interactions, control proteins involved in numerous biological functions. Retroviruses such as HIV-1 form by self-assembly when their genomic RNA (gRNA) traps Gag and GagPol polyprotein precursors. Infectivity requires extracellular budding of the particle followed by maturation, an ordered processing of approximately 2400 Gag and approximately 120 GagPol by the viral protease (PR). This leads to a condensed gRNA-NCp7 nucleocapsid and a CAp24-self-assembled capsid surrounding the RNP. The choreography by which all of these components dynamically interact during virus maturation is one of the missing milestones to fully depict the HIV life cycle. Here, we describe how HIV-1 has evolved a dynamic RNP granule with successive weak-strong-moderate quinary NC-gRNA networks during the sequential processing of the GagNC domain. We also reveal two palindromic RNA-binding triads on NC, KxxFxxQ and QxxFxxK, that provide quinary NC-gRNA interactions. Consequently, the nucleocapsid complex appears properly aggregated for capsid reassembly and reverse transcription, mandatory processes for viral infectivity. We show that PR is sequestered within this RNP and drives its maturation/condensation within minutes, this process being most effective at the end of budding. We anticipate such findings will stimulate further investigations of quinary interactions and emergent mechanisms in crowded environments throughout the wide and growing array of RNP granules.},

note = {1999-4915 (Electronic) 

1999-4915 (Linking) 

Journal Article},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{S.2020,

title = {Special Issue "Function and Structure of Viral Ribonucleoproteins Complexes"},

author = {S Bernacchi},

url = {https://www.mdpi.com/1999-4915/12/12/1355},

doi = { 10.3390/v12121355 },

year  = {2020},

date = {2020-11-26},

journal = {Viruses},

volume = {12},

number = {12},

pages = {E1355},

abstract = {RNA viruses are extraordinary evolution machines that efficiently ensure their replication by taking advantage of the association with viral and cellular components to form ribonucleic complexes (vRNPs) [...]. },

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{L.2020,

title = {Role of Purine-Rich Regions in Mason-Pfizer Monkey Virus (MPMV) Genomic RNA Packaging and Propagation },

author = {L M Ali and F N Pitchai and V Vivet-Boudou and A Chameettachal and A Jabeen and V N Pillai and F Mustafa and R Marquet and T A Rizvi

},

url = {https://www.frontiersin.org/articles/10.3389/fmicb.2020.595410/full},

doi = { 10.3389/fmicb.2020.595410 },

year  = {2020},

date = {2020-11-05},

journal = {Frontiers in Microbiology},

number = {11},

pages = {595410},

abstract = {A distinguishing feature of the Mason-Pfizer monkey virus (MPMV) packaging signal RNA secondary structure is a single-stranded purine-rich sequence (ssPurines) in close vicinity to a palindromic stem loop (Pal SL) that functions as MPMV dimerization initiation site (DIS). However, unlike other retroviruses, MPMV contains a partially base-paired repeat sequence of ssPurines (bpPurines) in the adjacent region. Both purine-rich sequences have earlier been proposed to act as potentially redundant Gag binding sites to initiate the process of MPMV genomic RNA (gRNA) packaging. The objective of this study was to investigate the biological significance of ssPurines and bpPurines in MPMV gRNA packaging by systematic mutational and biochemical probing analyses. Deletion of either ssPurines or bpPurines individually had no significant effect on MPMV gRNA packaging, but it was severely compromised when both sequences were deleted simultaneously. Selective 2' hydroxyl acylation analyzed by primer extension (SHAPE) analysis of the mutant RNAs revealed only mild effects on structure by deletion of either ssPurines or bpPurines, while the structure was dramatically affected by the two simultaneous deletions. This suggests that ssPurines and bpPurines play a redundant role in MPMV gRNA packaging, probably as Gag binding sites to facilitate gRNA capture and encapsidation. Interestingly, the deletion of bpPurines revealed an additional severe defect on RNA propagation that was independent of the presence or absence of ssPurines or the gRNA structure of the region. These findings further suggest that the bpPurines play an additional role in the early steps of MPMV replication cycle that is yet to be identified. },

keywords = {base paired purines, MARQUET, Mason-Pfizer monkey virus, PAILLART, retroviruses, RNA packaging, RNA secondary structure, RNA-Gag interaction, SHAPE, single-stranded purines, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Dynamic nanopore long-read sequencing analysis of HIV-1 splicing events during the early steps of infection},

author = {N Nguyen Quang and S Goudey and E Ségéral and A Mohammad and S Lemoine and C Blugeon and M Versapuech and J C Paillart and C Berlioz-Torrent and S Emiliani and S Gallois-Montbrun},

url = {https://pubmed.ncbi.nlm.nih.gov/32807178/},

doi = {10.1186/s12977-020-00533-1},

isbn = {32807178},

year  = {2020},

date = {2020-01-01},

journal = {Retrovirology},

volume = {17},

number = {1},

pages = {25},

abstract = {Background 

 

Alternative splicing is a key step in Human Immunodeficiency Virus type 1 (HIV-1) replication that is tightly regulated both temporally and spatially. More than 50 different transcripts can be generated from a single HIV-1 unspliced pre-messenger RNA (pre-mRNA) and a balanced proportion of unspliced and spliced transcripts is critical for the production of infectious virions. Understanding the mechanisms involved in the regulation of viral RNA is therefore of potential therapeutic interest. However, monitoring the regulation of alternative splicing events at a transcriptome-wide level during cell infection is challenging. Here we used the long-read cDNA sequencing developed by Oxford Nanopore Technologies (ONT) to explore in a quantitative manner the complexity of the HIV-1 transcriptome regulation in infected primary CD4+ T cells. 

Results 

 

ONT reads mapping to the viral genome proved sufficiently long to span all possible splice junctions, even distant ones, and to be assigned to a total of 150 exon combinations. Fifty-three viral RNA isoforms, including 14 new ones were further considered for quantification. Relative levels of viral RNAs determined by ONT sequencing showed a high degree of reproducibility, compared favourably to those produced in previous reports and highly correlated with quantitative PCR (qPCR) data. To get further insights into alternative splicing regulation, we then compiled quantifications of splice site (SS) usage and transcript levels to build ﾓsplice treesﾔ, a quantitative representation of the cascade of events leading to the different viral isoforms. This approach allowed visualizing the complete rewiring of SS usages upon perturbation of SS D2 and its impact on viral isoform levels. Furthermore, we produced the first dynamic picture of the cascade of events occurring between 12 and 24 h of viral infection. In particular, our data highlighted the importance of non-coding exons in viral RNA transcriptome regulation. 

Conclusion 

 

ONT sequencing is a convenient and reliable strategy that enabled us to grasp the dynamic of the early splicing events modulating the viral RNA landscape in HIV-1 infected cells. 

Background},

keywords = {HIV RNA Alternative splicing Viral transcriptome ONT long-read sequencing, MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Zinc Fingers in HIV-1 Gag Precursor Are Not Equivalent for gRNA Recruitment at the Plasma Membrane},

author = {E Boutant and J Bonzi and H Anton and M Bin Nasim and R Cathagne and E Real and D Dujardin and P Carl and P Didier and J C Paillart and R Marquet and Y Mely and H de Rocquigny and S Bernacchi},

url = {https://pubmed.ncbi.nlm.nih.gov/32574557/},

doi = {10.1016/j.bpj.2020.05.035},

isbn = {32574557},

year  = {2020},

date = {2020-01-01},

journal = {Biophys J},

volume = {119},

number = {2},

pages = {419-433},

abstract = {The human immunodeficiency virus type 1 Gag precursor specifically selects the unspliced viral genomic RNA (gRNA) from the bulk of cellular and spliced viral RNAs via its nucleocapsid (NC) domain and drives gRNA encapsidation at the plasma membrane (PM). To further identify the determinants governing the intracellular trafficking of Gag-gRNA complexes and their accumulation at the PM, we compared, in living and fixed cells, the interactions between gRNA and wild-type Gag or Gag mutants carrying deletions in NC zinc fingers (ZFs) or a nonmyristoylated version of Gag. Our data showed that the deletion of both ZFs simultaneously or the complete NC domain completely abolished intracytoplasmic Gag-gRNA interactions. Deletion of either ZF delayed the delivery of gRNA to the PM but did not prevent Gag-gRNA interactions in the cytoplasm, indicating that the two ZFs display redundant roles in this respect. However, ZF2 played a more prominent role than ZF1 in the accumulation of the ribonucleoprotein complexes at the PM. Finally, the myristate group, which is mandatory for anchoring the complexes at the PM, was found to be dispensable for the association of Gag with the gRNA in the cytosol.},

keywords = {MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{Verriez2020,

title = {Les APOBEC : histoire d’une famille de protéines antivirales et mutagènes.},

author = {C Verriez, R Marquet, J-C Paillart and B Stupfler},

year  = {2020},

date = {2020-01-01},

journal = {Virologie},

volume = {24},

number = {6},

pages = {381-418},

abstract = {La réponse immunitaire innée est une réponse non spécifique qui constitue la première ligne de défense en cas d’infection, notamment en permettant l’élimination des pathogènes par phagocytose ou apoptose. Au sein des cellules immunitaires, cette réponse innée se caractérise entre autres par la synthèse de protéines nommées facteurs de restriction dont le rôle est d’inhiber la réplication virale. Parmi ces facteurs, les protéines de la famille APOBEC3 (Apolipoprotein B mRNA-editing Enzyme Catalytic polypeptide-like 3 ou A3) constituent des facteurs antiviraux majeurs qui ciblent de nombreux types de virus. L’une des cibles des A3 est le virus de l’immunodéficience humaine de type 1 (VIH-1) : l’activité désaminase de certaines A3 convertit une fraction des cytidines du génome viral en uridines et perturbe son expression. Néanmoins, le VIH-1 contrecarre les A3 en exprimant la protéine Vif qui les inhibe en détournant divers mécanismes cellulaires. Par ailleurs, les APOBEC3 participent au maintien de l’intégrité génétique par l’inhibition des rétroéléments mais contribuent également à la cancérogenèse, à l’image d’A3A et A3B, deux facteurs majeurs dans ce processus. L’éventail de leurs activités, combiné aux récentes études montrant leur implication dans la régulation de virus émergents (Zika, SARS-CoV-2), permettent d’envisager les A3 ainsi que leurs partenaires viraux comme axes thérapeutiques.},

keywords = {APOBEC3, cancer, Facteurs de restriction, MARQUET, PAILLART, Unité ARN, vif, VIH-1},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {The evolution of RNA structural probing methods: From gels to next-generation sequencing},

author = {E Mailler and J C Paillart and R Marquet and R P Smyth and V Vivet-Boudou},

url = {https://www.ncbi.nlm.nih.gov/pubmed/30485688?dopt=Abstract},

doi = {10.1002/wrna.1518},

isbn = {30485688},

year  = {2019},

date = {2019-01-01},

journal = {Wiley Interdiscip Rev RNA},

volume = {10},

number = {2},

pages = {e1518},

abstract = {RNA molecules are important players in all domains of life and the study of the relationship between their multiple flexible states and the associated biological roles has increased in recent years. For several decades, chemical and enzymatic structural probing experiments have been used to determine RNA structure. During this time, there has been a steady improvement in probing reagents and experimental methods, and today the structural biologist community has a large range of tools at its disposal to probe the secondary structure of RNAs in vitro and in cells. Early experiments used radioactive labeling and polyacrylamide gel electrophoresis as read-out methods. This was superseded by capillary electrophoresis, and more recently by next-generation sequencing. Today, powerful structural probing methods can characterize RNA structure on a genome-wide scale. In this review, we will provide an overview of RNA structural probing methodologies from a historical and technical perspective. This article is categorized under: RNA Structure and Dynamics > RNA Structure, Dynamics, and Chemistry RNA Methods > RNA Analyses in vitro and In Silico RNA Methods > RNA Analyses in Cells.},

keywords = {MARQUET, NGS RNA probing structure, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Purification and Functional Characterization of a Biologically Active Full-Length Feline Immunodeficiency Virus (FIV) Pr50^{Gag}.},

author = {A Krishnan and V Pillai and A Chameettachal and L M Ali and F Nuzra Nagoor Pitchai and S Tariq and F Mustafa and R Marquet and T A Rizvi},

url = {https://www.ncbi.nlm.nih.gov/pubmed/31357656?report=&dispmax=200&tool=PubCrawler_2.23},

doi = {10.3390/v11080689},

isbn = {31357656},

year  = {2019},

date = {2019-01-01},

journal = {Viruses},

volume = {11},

number = {8},

pages = {689},

abstract = {The feline immunodeficiency virus (FIV) full-length Pr50Gag precursor is a key player in the assembly of new viral particles. It is also a critical component of the efficient selection and packaging of two copies of genomic RNA (gRNA) into the newly formed virus particles from a wide pool of cellular and spliced viral RNA. To understand the molecular mechanisms involved during FIV gRNA packaging, we expressed the His6-tagged and untagged recombinant FIV Pr50Gag protein both in eukaryotic and prokaryotic cells. The recombinant Pr50Gag-His6-tag fusion protein was purified from soluble fractions of prokaryotic cultures using immobilized metal affinity chromatography (IMAC). This purified protein was able to assemble in vitro into virus-like particles (VLPs), indicating that it preserved its ability to oligomerize/multimerize. Furthermore, VLPs formed in eukaryotic cells by the FIV full-length Pr50Gag both in the presence and absence of His6-tag could package FIV sub-genomic RNA to similar levels, suggesting that the biological activity of the recombinant full-length Pr50Gag fusion protein was retained in the presence of His6-tag at the carboxy terminus. Successful expression and purification of a biologically active, recombinant full-length Pr50Gag-His6-tag fusion protein will allow study of the intricate RNA-protein interactions involved during FIV gRNA encapsidation.},

keywords = {Gag protein purification His-tag fusion protein Pr50Gag protein expression feline immunodeficiency virus (FIV) retroviral RNA packaging viral assembly, MARQUET, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Stabilizing role of structural elements within the 5´ Untranslated Region (UTR) and gag sequences in Mason-Pfizer monkey virus (MPMV) genomic RNA packaging},

author = {R M Kalloush and V Vivet-Boudou and L M Ali and V Pillai and F Mustafa and R Marquet and T A Rizvi},

url = {https://www.ncbi.nlm.nih.gov/pubmed/30773097?dopt=Abstract},

doi = {10.1080/15476286.2019.1572424},

isbn = {30773097},

year  = {2019},

date = {2019-01-01},

journal = {RNA Biol},

volume = {16},

number = {5},

pages = {612-625},

abstract = {The Mason-Pfizer monkey virus (MPMV) genomic RNA (gRNA) packaging signal is a highly-structured element with several stem-loops held together by two phylogenetically conserved long-range interactions (LRIs) between U5 and gag complementary sequences. These LRIs play a critical role in maintaining the structure of the 5´ end of the MPMV gRNA. Thus, one could hypothesize that the overall RNA secondary structure of this region is further architecturally held together by three other stem loops (SL3, Gag SL1, and Gag SL2) comprising of sequences from the distal parts of the 5´untranslated region (5' UTR) to ~ 120 nucleotides into gag, excluding gag sequences involved in forming the U5-Gag LRIs. To provide functional evidence for the biological significance of these stem loops during gRNA encapsidation, these structural motifs were mutated and their effects on MPMV RNA packaging and propagation were tested in a single round trans-complementation assay. The mutant RNA structures were further studied by high throughput SHAPE (hSHAPE) assay. Our results reveal that sequences involved in forming these three stem loops do not play crucial roles at an individual level during MPMV gRNA packaging or propagation. Further structure-function analysis indicates that the U5-Gag LRIs have a more important architectural role in stabilizing the higher order structure of the 5´ UTR than the three stem loops which have a more secondary and perhaps indirect role in stabilizing the overall RNA secondary structure of the region. Our work provides a better understanding of the molecular interactions that take place during MPMV gRNA packaging.},

keywords = {MARQUET, MARQUET  PAILLART  Mason-Pfizer monkey virus (MPMV)  RNA packaging  RNA secondary structure  Retroviruses  U5/Gag LRIs  gag, Mason-Pfizer monkey virus (MPMV) RNA packaging RNA secondary structure Retroviruses U5/Gag LRIs gag, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}