Pathogenesis of bacterial infections and immunity

@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/s12977-023-00627-6},

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{pmid36996029,

title = {A snapshot on HIV-1 evolution through the identification of phylogenetic-specific properties of HIV-1 integrases M/O},

author = {Elenia Toccafondi and Marine Kanja and Flore Winter and Daniela Lener and Matteo Negroni},

doi = {10.1371/journal.ppat.1011207},

issn = {1553-7374},

year  = {2023},

date = {2023-03-01},

urldate = {2023-03-01},

journal = {PLoS Pathog},

volume = {19},

number = {3},

pages = {e1011207},

abstract = {Transmissions of simian viruses to humans has originated the different groups of HIV-1. We recently identified a functional motif (CLA), in the C-terminal domain of the integrase, essential for integration in HIV-1 group M. Here, we found that the motif is instead dispensable in group O isolates, because of the presence, in the N-terminal domain of HIV-1 O of a specific sequence, Q7G27P41H44, that we define as the NOG motif. Alterations of reverse transcription and of 3' processing observed by mutating the CLA motif of IN M are fully rescued to wt levels by inserting the sequence of the NOG motif in the N-ter of the protein. These results indicate that the two motifs (CLA and NOG) functionally complement each other and a working model accounting for these observations is proposed. The establishment of these two alternative motifs seems to be due to the different phylogenetic origin and history of these two groups. Indeed, the NOG motif is already present in the ancestor of group O (SIVgor) while it is absent from SIVcpzPtt, the ancestor of group M. The CLA motif, instead, seems to have emerged after SIVcpzPtt has been transferred to humans, since no conservation is found at the same positions in these simian viruses. These results show the existence of two-group specific motifs in HIV-1 M and O integrases. In each group, only one of the motifs is functional, potentially leading the other motif to diverge from its original function and, in an evolutionary perspective, assist other functions of the protein, further increasing HIV genetic diversity.},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {CDX2 regulates ACE expression in blood development and leukemia cells},

author = {R El Omar and E Julien and K Biasch and B Guffroy and B Lioure and L Vallat and I Gross and C Domon-Dell and F Lanza and C Gachet and M Negroni and J N Freund and M Tavian},

url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33843985},

doi = {10.1182/bloodadvances.2020003563},

isbn = {33843985},

year  = {2021},

date = {2021-01-01},

journal = {Blood Adv},

volume = {5},

number = {7},

pages = {2012-2016},

note = {2473-9537 (Electronic) 

2473-9529 (Linking) 

Journal Article},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {HIV-1 Capsid Core: A Bullet to the Heart of the Target Cell},

author = {E Toccafondi and D Lener and M Negroni},

url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33868211},

doi = {10.3389/fmicb.2021.652486},

isbn = {33868211},

year  = {2021},

date = {2021-01-01},

journal = {Front Microbiol},

volume = {12},

pages = {652486},

abstract = {The first step of the intracellular phase of retroviral infection is the release of the viral capsid core in the cytoplasm. This structure contains the viral genetic material that will be reverse transcribed and integrated into the genome of infected cells. Up to recent times, the role of the capsid core was considered essentially to protect this genetic material during the earlier phases of this process. However, increasing evidence demonstrates that the permanence inside the cell of the capsid as an intact, or almost intact, structure is longer than thought. This suggests its involvement in more aspects of the infectious cycle than previously foreseen, particularly in the steps of viral genomic material translocation into the nucleus and in the phases preceding integration. During the trip across the infected cell, many host factors are brought to interact with the capsid, some possessing antiviral properties, others, serving as viral cofactors. All these interactions rely on the properties of the unique component of the capsid core, the capsid protein CA. Likely, the drawback of ensuring these multiple functions is the extreme genetic fragility that has been shown to characterize this protein. Here, we recapitulate the busy agenda of an HIV-1 capsid in the infectious process, in particular in the light of the most recent findings.},

note = {1664-302X (Print) 

1664-302X (Linking) 

Journal Article 

Review},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{nokey,

title = {The HIV-1 Integrase C-Terminal domain induces TAR RNA structural changes promoting Tat binding},

author = {C. Rocchi and C. Louvat and A. Miele and J. Batisse and C. Guillon and L. Ballut and D. Lener and M. Negroni and M. Ruff and P. Gouet and F. Fiorini},

url = {https://www.biorxiv.org/content/10.1101/2021.10.21.465253v1.full.pdf+html},

doi = {10.1101/2021.10.21.465253},

year  = {2021},

date = {2021-01-01},

abstract = {Recent evidence indicated that HIV-1 Integrase (IN) binds genomic viral RNA (gRNA) playing a critical role in viral particle morphogenesis and gRNA stability in host cells. Combining biophysical and biochemical approaches we show that the C-terminal flexible 18-residues tail of IN acts as a sensor of the peculiar apical structure of trans-activation 

 response element RNA (TAR), directly interacting with its hexaloop. We highlighted how the whole IN C-terminal domain, once bound to TAR, can change its structure assisting the binding of Tat, the HIV trans-activator protein, which finally displaces IN from TAR. Our results are consistent with the emerging role of IN in early stage of proviral transcription and suggest new steps of HIV-1 life cycle that can be considered as therapeutic targets.},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{Duvergé2020,

title = {Pseudotyping Lentiviral Vectors: When the Clothes Make the Virus },

author = {A Duvergé and M Negroni},

url = {https://doi.org/10.3390/v12111311},

doi = {10.3390/v12111311},

year  = {2020},

date = {2020-11-11},

journal = {Viruses},

volume = {12},

number = {11},

pages = {1311},

abstract = {Delivering transgenes to human cells through transduction with viral vectors constitutes one of the most encouraging approaches in gene therapy. Lentivirus-derived vectors are among the most promising vectors for these approaches. When the genetic modification of the cell must be performed in vivo, efficient specific transduction of the cell targetsof the therapy in the absence of off-targeting constitutes the Holy Grail of gene therapy. For viral therapy, this is largely determined by the characteristics of the surface proteins carried by the vector. In this regard, an important property of lentiviral vectors is the possibility of being pseudotyped by envelopes of other viruses, widening the panel of proteins with which they can bearmed. Here, we discuss how this is achieved at  the  molecular  level  and  what  the  properties  and  the  potentialities  of  the  different  envelope proteins that can be used for pseudotyping these vectorsare.},

keywords = {envelope proteins, gene therapy, lentiviral vectors, NEGRONI, pseudotyping, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {NKNK: a New Essential Motif in the C-Terminal Domain of HIV-1 Group M Integrases},

author = {M Kanja and P Cappy and N Levy and O Oladosu and S Schmidt and P Rossolillo and F Winter and R Gasser and C Moog and M Ruff and M Negroni and D Lener},

url = {https://pubmed.ncbi.nlm.nih.gov/32727879/},

doi = {10.1128/JVI.01035-20},

isbn = {32727879},

year  = {2020},

date = {2020-01-01},

journal = {J Virol},

volume = {94},

number = {20},

pages = {e01035-01020},

abstract = {Using coevolution network interference based on comparison of two phylogenetically distantly related isolates, one from the main group M and the other from the minor group O of HIV-1, we identify, in the C-terminal domain (CTD) of integrase, a new functional motif constituted by four noncontiguous amino acids (N222K240N254K273). Mutating the lysines abolishes integration through decreased 3' processing and inefficient nuclear import of reverse-transcribed genomes. Solution of the crystal structures of wild-type (wt) and mutated CTDs shows that the motif generates a positive surface potential that is important for integration. The number of charges in the motif appears more crucial than their position within the motif. Indeed, the positions of the K's could be permutated or additional K's could be inserted in the motif, generally without affecting integration per se Despite this potential genetic flexibility, the NKNK arrangement is strictly conserved in natural sequences, indicative of an effective purifying selection exerted at steps other than integration. Accordingly, reverse transcription was reduced even in the mutants that retained wt integration levels, indicating that specifically the wt sequence is optimal for carrying out the multiple functions that integrase exerts. We propose that the existence of several amino acid arrangements within the motif, with comparable efficiencies of integration per se, might have constituted an asset for the acquisition of additional functions during viral evolution.IMPORTANCE Intensive studies of HIV-1 have revealed its extraordinary ability to adapt to environmental and immunological challenges, an ability that is also at the basis of antiviral treatment escape. Here, by deconvoluting the different roles of the viral integrase in the various steps of the infectious cycle, we report how the existence of alternative equally efficient structural arrangements for carrying out one function opens up the possibility of adapting to the optimization of further functionalities exerted by the same protein. Such a property provides an asset to increase the efficiency of the infectious process. On the other hand, though, the identification of this new motif provides a potential target for interfering simultaneously with multiple functions of the protein.},

keywords = {evolution human immunodeficiency virus integrase phylogenetic groups, NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {RNA Structure-A Neglected Puppet Master for the Evolution of Virus and Host Immunity},

author = {R P Smyth and M Negroni and A M Lever and J Mak and J C Kenyon},

url = {https://www.ncbi.nlm.nih.gov/pubmed/30283444?dopt=Abstract},

doi = {10.3389/fimmu.2018.02097},

isbn = {30283444},

year  = {2018},

date = {2018-01-01},

journal = {Front Immunol},

volume = {9},

pages = {2097},

abstract = {The central dogma of molecular biology describes the flow of genetic information from DNA to protein via an RNA intermediate. For many years, RNA has been considered simply as a messenger relaying information between DNA and proteins. Recent advances in next generation sequencing technology, bioinformatics, and non-coding RNA biology have highlighted the many important roles of RNA in virtually every biological process. Our understanding of RNA biology has been further enriched by a number of significant advances in probing RNA structures. It is now appreciated that many cellular and viral biological processes are highly dependent on specific RNA structures and/or sequences, and such reliance will undoubtedly impact on the evolution of both hosts and viruses. As a contribution to this special issue on host immunity and virus evolution, it is timely to consider how RNA sequences and structures could directly influence the co-evolution between hosts and viruses. In this manuscript, we begin by stating some of the basic principles of RNA structures, followed by describing some of the critical RNA structures in both viruses and hosts. More importantly, we highlight a number of available new tools to predict and to evaluate novel RNA structures, pointing out some of the limitations readers should be aware of in their own analyses.},

keywords = {NEGRONI, RNA structure immune evasion secondary structure viral RNA viral evolution, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {HIV-1 sequences in the epidemic suggest an alternative pathway for the generation of the Long Terminal Repeats},

author = {P Cappy and A Moisan and F De Oliveira and J C Plantier and M Negroni},

url = {https://www.nature.com/articles/s41598-017-14135-z},

doi = {10.1038/s41598-017-14135-z},

isbn = {29057964},

year  = {2017},

date = {2017-01-01},

journal = {Sci Rep},

volume = {7},

number = {1},

pages = {13715},

abstract = {To generate the long-terminal repeats (LTR) that border the integrated viral genome, two-strand transfer steps must occur during reverse transcription. Analysis of the genetic polymorphisms that are present in the LTR of HIV-1 heterozygous virions in single infection cycle studies has revealed which of the two copies of genomic RNAs is used for each transfer event. Thus, the first event of strand transfer has been described to be either intra- or intermolecular, while the second event is generally intramolecular. Here, we repeated these analyses using sequences from HIV databases and extended the study to the regions surrounding the LTR. We observed a striking correlation between the pattern of recombination in the LTR and the phylogenetic origin of the surrounding sequences. This correlation suggests that the second-strand transfer can be either intra- or intermolecular and, interestingly, could reflect an effect of proximity between nucleic acids that would guide this transfer. This factor could be particularly relevant for heterozygous viruses containing highly divergent genomic RNAs, such as those considered in the present study.},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {A step forward understanding HIV-1 diversity},

author = {R P Smyth and M Negroni},

url = {http://www.ncbi.nlm.nih.gov/pubmed/27093884?dopt=Abstract},

doi = {10.1186/s12977-016-0259-8},

isbn = {27093884},

year  = {2016},

date = {2016-01-01},

journal = {Retrovirology},

volume = {13},

number = {1},

pages = {27},

abstract = {Human immunodeficiency virus (HIV) populations are characterized by extensive genetic diversity. Antigenic diversification is essential for escape from immune selection and therapy, and remains one of the major obstacles for the development of an efficient vaccine strategy. Even if intensive efforts have been made for understanding the molecular mechanisms responsible for genetic diversity in HIV, conclusive data in vivo is still lacking. Recent works have addressed this issue, focusing on the identification of the sources of genetic diversity during in vivo infections and on the estimate of the pervasiveness of genetic recombination during replication in vivo. Surprisingly, it appears that despite the error-prone nature of the viral polymerase, the bulk of mutations found in patients are indeed due to the effect of a cellular restriction factor. This factor tends to hypermutate the viral genome abolishing viral infectivity. When hypermutation is incomplete, the virus retains infectivity and converts the effect of the cellular factor to its advantage by exploiting it to generate genetic diversity that is beneficial for viral propagation. This view contrasts the long-standing dogma that viral diversity is due to the intrinsic error-prone nature of the viral replication cycle. Besides hypermutations and mutations, recombination is also a pervasive source of genetic diversity. The estimate of the frequency at which this process takes place in vivo has remained elusive, despite extensive efforts in this sense. Now, using single genome amplification, and starting from publically available datasets, it has been obtained a confirmation of the estimates previously made using tissue culture studies. These recent findings are presented here and their implications for the development of future researches are discussed.},

keywords = {alternative splicing aminoacyl-tRNA synthetase enzyme kinetics mitochondria mitochondrial disease threonyl-tRNA synthetase, MARQUET, NEGRONI, PAILLART, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Buffering deleterious polymorphisms in highly constrained parts of HIV-1 envelope by flexible regions.},

author = {R Gasser and M Hamoudi and M Pellicciotta and Z Zhou and C Visdeloup and P Colin and M Braibant and B Lagane and M Negroni},

url = {http://www.ncbi.nlm.nih.gov/pubmed/27473399?dopt=Abstract},

doi = {10.1186/s12977-016-0285-6},

year  = {2016},

date = {2016-01-01},

urldate = {2016-01-01},

journal = {Retrovirology},

volume = {13},

number = {1},

pages = {50},

abstract = {BACKGROUND: 

 

Covariation is an essential process that leads to coevolution of parts of proteins and genomes. In organisms subject to strong selective pressure, coevolution is central to keep the balance between the opposite requirements of antigenic variation and retention of functionality. Being the viral component most exposed to the external environment, the HIV-1 glycoprotein gp120 constitutes the main target of the immune response. Accordingly its more external portions are characterised by extensive sequence heterogeneity fostering constant antigenic variation. 

RESULTS: 

 

We report that a single polymorphism, present at the level of the viral population in the conserved internal region C2, was sufficient to totally abolish Env functionality when introduced in an exogenous genetic context. The prominent defect of the non-functional protein is a block occurring after recognition of the co-receptor CCR5, likely due to an interference with the subsequent conformational changes that lead to membrane fusion. We also report that the presence of compensatory polymorphisms at the level of the external and hypervariable region V3 fully restored the functionality of the protein. The functional revertant presents different antigenic profiles and sensitivity to the entry inhibitor TAK 779. 

CONCLUSIONS: 

 

Our data suggest that variable regions, besides harbouring intrinsic extensive antigenic diversity, can also contribute to sequence diversification in more structurally constrained parts of the gp120 by buffering the deleterious effect of polymorphisms, further increasing the genetic flexibility of the protein and the antigenic repertoire of the viral population.},

keywords = {Antigenic variation Coevolution Envelope HIV Viral entry, NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {The external domains of the HIV-1 envelope are a mutational cold spot.},

author = {R Geller and P Domingo-Calap and J M Cuevas and P Rossolillo and M Negroni and R Sanjuán},

url = {http://www.ncbi.nlm.nih.gov/pubmed/26450412?dopt=Abstract},

doi = {10.1038/ncomms9571},

isbn = {26450412},

year  = {2015},

date = {2015-01-01},

journal = {Nat Commun},

volume = {6},

pages = {8571},

abstract = {In RNA viruses, mutations occur fast and have large fitness effects. While this affords remarkable adaptability, it can also endanger viral survival due to the accumulation of deleterious mutations. How RNA viruses reconcile these two opposed facets of mutation is still unknown. Here we show that, in human immunodeficiency virus (HIV-1), spontaneous mutations are not randomly located along the viral genome. We find that the viral mutation rate experiences a threefold reduction in the region encoding the most external domains of the viral envelope, which are strongly targeted by neutralizing antibodies. This contrasts with the hypermutation mechanisms deployed by other, more slowly mutating pathogens such as DNA viruses and bacteria, in response to immune pressure. We show that downregulation of the mutation rate in HIV-1 is exerted by the template RNA through changes in sequence context and secondary structure, which control the activity of apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (A3)-mediated cytidine deamination and the fidelity of the viral reverse transcriptase.},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Potent Sensitisation of Cancer Cells to Anticancer Drugs by a Quadruple Mutant of the Human Deoxycytidine Kinase.},

author = {S T Coulibaly and P Rossolillo and F Winter and F K Kretzschmar and M Brayé and D P Martin and D Lener and M Negroni},

url = {http://www.ncbi.nlm.nih.gov/pubmed/26485161?dopt=Abstract},

doi = {10.1371/journal.pone.0140741},

isbn = {26485161},

year  = {2015},

date = {2015-01-01},

journal = {PLoS One},

volume = {10},

number = {10},

pages = {e0140741},

abstract = {Identifying enzymes that, once introduced in cancer cells, lead to an increased efficiency of treatment constitutes an important goal for biomedical applications. Using an original procedure whereby mutant genes are generated based on the use of conditional lentivector genome mobilisation, we recently described, for the first time, the identification of a human deoxycytidine kinase (dCK) mutant (G12) that sensitises a panel of cancer cell lines to treatment with the dCK analogue gemcitabine. Here, starting from the G12 variant itself, we generated a new library and identified a mutant (M36) that triggers even greater sensitisation to gemcitabine than G12. With respect to G12, M36 presents an additional mutation located in the region that constitutes the interface of the dCK dimer. The simple presence of this mutation halves both the IC50 and the proportion of residual cells resistant to the treatment. Furthermore, the use of vectors with self-inactivating LTRs leads to an increased sensitivity to treatment, a result compatible with a relief of the transcriptional interference exerted by the U3 promoter on the internal promoter that drives the expression of M36. Importantly, a remarkable effect is also observed in treatments with the anticancer compound cytarabine (AraC), for which a 10,000 fold decrease in IC50 occurred. By triggering the sensitisation of various cancer cell types with poor prognosis to two commonly used anticancer compounds M36 is a promising candidate for suicide gene approaches.},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}

@article{,

title = {Retrovolution: HIV-Driven Evolution of Cellular Genes and Improvement of Anticancer Drug Activation},

author = {P Rossolillo and F Winter and E Simon-Loriere and S Gallois-Montbrun and M Negroni},

url = {http://www.plosgenetics.org/article/info%3Adoi%2F10.1371%2Fjournal.pgen.1002904},

doi = {10.1371/journal.pgen.1002904},

isbn = {22927829},

year  = {2012},

date = {2012-01-01},

journal = {PLoS Genet},

volume = {8},

number = {8},

pages = {e1002904},

abstract = {In evolution strategies aimed at isolating molecules with new functions, screening for the desired phenotype is generally performed in vitro or in bacteria. When the final goal of the strategy is the modification of the human cell, the mutants selected with these preliminary screenings may fail to confer the desired phenotype, due to the complex networks that regulate gene expression in higher eukaryotes. We developed a system where, by mimicking successive infection cycles with HIV-1 derived vectors containing the gene target of the evolution in their genome, libraries of gene mutants are generated in the human cell, where they can be directly screened. As a proof of concept we created a library of mutants of the human deoxycytidine kinase (dCK) gene, involved in the activation of nucleoside analogues used in cancer treatment, with the aim of isolating a variant sensitizing cancer cells to the chemotherapy compound Gemcitabine, to be used in gene therapy for anti-cancer approaches or as a poorly immunogenic negative selection marker for cell transplantation approaches. We describe the isolation of a dCK mutant, G12, inducing a 300-fold sensitization to Gemcitabine in cells originally resistant to the prodrug (Messa 10K), an effect 60 times stronger than the one induced by the wt enzyme. The phenotype is observed in different tumour cell lines irrespective of the insertion site of the transgene and is due to a change in specificity of the mutated kinase in favour of the nucleoside analogue. The mutations characterizing G12 are distant from the active site of the enzyme and are unpredictable on a rational basis, fully validating the pragmatic approach followed. Besides the potential interest of the G12 dCK variant for therapeutic purposes, the methodology developed is of interest for a large panel of applications in biotechnology and basic research.},

keywords = {NEGRONI, Unité ARN},

pubstate = {published},

tppubtype = {article}

}