Retroviral integration site selection: a running Gag?

The ability of retroviruses to integrate their genomes into host chromatin is a key step for the completion of their replication cycle. Selection of a suitable chromosomal integration site has been described as a hierarchical mechanism involving both cellular and viral proteins but the exact molecular determinants are still unclear. We recently showed that the spumaretrovirus prototype foamy virus (PFV) Gag protein is acting as a chromatin tether by interacting with the nucleosome acidic patch (Lesbats et al. PNAS 114(21)). Disruption of the nucleosome binding leads to a dramatic delocalization of both the viral particles and the integration sites accompanied with a reduction of integrated genes expression. These data show for the first time a direct interaction between retroviral structural proteins with the host chromosomes, and highlight their importance in the integration sites selection

Targeting the Nucleosome Acidic Patch by Viral Proteins: Two Birds with One Stone?

The past decade illuminated the H2A-H2B acidic patch as a cornerstone for both nucleosome recognition and chromatin structure regulation. Higher-order folding of chromatin arrays is mediated by interactions of histone H4 tail with an adjacent nucleosome acidic patch. Dynamic chromatin folding ensures a proper regulation of nuclear functions fundamental to cellular homeostasis. Many cellular factors have been shown to act on chromatin by tethering nucleosomes via an arginine anchor binding to the acidic patch. This tethering mechanism has also been described for several viral proteins. In this minireview, we will discuss the structural basis for acidic patch engagement by viral proteins and the implications during respective viral infections. We will also discuss a model in which acidic patch occupancy by these non-self viral proteins alters the local chromatin state by preventing H4 tail-mediated higher-order chromatin folding

Structural basis for spumavirus GAG tethering to chromatin

The interactions between a retrovirus and host cell chromatin that underlie integration and provirus expression are poorly understood. The prototype foamy virus (PFV) structural protein GAG associates with chromosomes via a chromatin-binding sequence (CBS) located within its C-terminal region. Here, we show that the PFV CBS is essential and sufficient for a direct interaction with nucleosomes and present a crystal structure of the CBS bound to a mononucleosome. The CBS interacts with the histone octamer, engaging the H2A–H2B acidic patch in a manner similar to other acidic patch-binding proteins such as herpesvirus latency-associated nuclear antigen (LANA). Substitutions of the invariant arginine anchor residue in GAG result in global redistribution of PFV and macaque simian foamy virus (SFVmac) integration sites toward centromeres, dampening the resulting proviral expression without affecting the overall efficiency of integration. Our findings underscore the importance of retroviral structural proteins for integration site selection and the avoidance of genomic junkyards

Functional architecture of the hiv-1 integration complex

L’intégrase (IN) du VIH-1 est une enzyme clé catalysant l’insertion de l’ADN proviral dans le génome cellulaire au sein d’un complexe nucléoprotéique d’intégration.Les nombreux efforts apportés à l’étude du mécanisme d’intégration ont permis d’accumuler de multiples données sur ce processus complexe. Cependant plusieurs questions essentielles demeurent sans réponses en particulier concernant l’architecture fonctionnelle des complexes d’intégration de VIH-1. En effet même si les complexes actifs pour l’intégration sont relativement bien définis leur chronologie précoce de formation demeure inconnue. De même les phases tardives de leur association au substrat naturel d’intégration que constitue la chromatine restent floues. Enfin le rôle des facteurs du complexe de préintégration (CPI) dans la régulation des ces mécanismes doit être déterminé.Mon travail de thèse s’est reposé sur trois axes s’articulant autour de ces questions:-Comment est régulée la dynamique des phases précoces d’attachement des oligomères d’intégrase sur les extrémités virales ?-Quel est l’impact de la structure de l’ADN cible et de la chromatine sur l’association active des complexes d’intégration ?-Quel(s) rôle(s) joue(nt) les facteurs du CPI dans ces phases ?HIV-1 integrase (IN) is a key enzyme catalyzing the proviral DNA insertion into the cellular genome within a nucleoprotein integration complex.The numerous efforts made on the mechanism of integration have led to the accumulation of substantial data on this process. However many important questions are still open particularly on the functional architecture of the HIV-1 integration complexes. Indeed even if the active complexes involved in the catalysis of the integration are well described, the chronology of their early formation is still unknown. Moreover, the late association of these complexes with the host chromatin is also obscure. Finally the involvement of the IN cofactors inside the preintegration complex on these steps has to be elucidated.The project of my PhD relied on three axis articulated on these questions:-What is the dynamic of the IN oligomers association on the DNA viral ends?-What is the impact of the target DNA chromatin structure on the functional association of the integration complexes?-Involvement of the PIC factors on these steps

Functional architecture of the hiv-1 integration complex

L’intégrase (IN) du VIH-1 est une enzyme clé catalysant l’insertion de l’ADN proviral dans le génome cellulaire au sein d’un complexe nucléoprotéique d’intégration.Les nombreux efforts apportés à l’étude du mécanisme d’intégration ont permis d’accumuler de multiples données sur ce processus complexe. Cependant plusieurs questions essentielles demeurent sans réponses en particulier concernant l’architecture fonctionnelle des complexes d’intégration de VIH-1. En effet même si les complexes actifs pour l’intégration sont relativement bien définis leur chronologie précoce de formation demeure inconnue. De même les phases tardives de leur association au substrat naturel d’intégration que constitue la chromatine restent floues. Enfin le rôle des facteurs du complexe de préintégration (CPI) dans la régulation des ces mécanismes doit être déterminé.Mon travail de thèse s’est reposé sur trois axes s’articulant autour de ces questions:-Comment est régulée la dynamique des phases précoces d’attachement des oligomères d’intégrase sur les extrémités virales ?-Quel est l’impact de la structure de l’ADN cible et de la chromatine sur l’association active des complexes d’intégration ?-Quel(s) rôle(s) joue(nt) les facteurs du CPI dans ces phases ?HIV-1 integrase (IN) is a key enzyme catalyzing the proviral DNA insertion into the cellular genome within a nucleoprotein integration complex.The numerous efforts made on the mechanism of integration have led to the accumulation of substantial data on this process. However many important questions are still open particularly on the functional architecture of the HIV-1 integration complexes. Indeed even if the active complexes involved in the catalysis of the integration are well described, the chronology of their early formation is still unknown. Moreover, the late association of these complexes with the host chromatin is also obscure. Finally the involvement of the IN cofactors inside the preintegration complex on these steps has to be elucidated.The project of my PhD relied on three axis articulated on these questions:-What is the dynamic of the IN oligomers association on the DNA viral ends?-What is the impact of the target DNA chromatin structure on the functional association of the integration complexes?-Involvement of the PIC factors on these steps

Chem Rev

The integration of a DNA copy of the viral RNA genome into host chromatin is the defining step of retroviral replication. This enzymatic process is catalyzed by the virus-encoded integrase protein, which is conserved among retroviruses and LTR-retrotransposons. Retroviral integration proceeds via two integrase activities: 3'-processing of the viral DNA ends, followed by the strand transfer of the processed ends into host cell chromosomal DNA. Herein we review the molecular mechanism of retroviral DNA integration, with an emphasis on reaction chemistries and architectures of the nucleoprotein complexes involved. We additionally discuss the latest advances on anti-integrase drug development for the treatment of AIDS and the utility of integrating retroviral vectors in gene therapy applications

Retroviral DNA Integration

The integration of a DNA copy of the viral RNA genome into host chromatin is the defining step of retroviral replication. This enzymatic process is catalyzed by the virus-encoded integrase protein, which is conserved among retroviruses and LTR-retrotransposons. Retroviral integration proceeds via two integrase activities: 3′-processing of the viral DNA ends, followed by the strand transfer of the processed ends into host cell chromosomal DNA. Herein we review the molecular mechanism of retroviral DNA integration, with an emphasis on reaction chemistries and architectures of the nucleoprotein complexes involved. We additionally discuss the latest advances on anti-integrase drug development for the treatment of AIDS and the utility of integrating retroviral vectors in gene therapy applications

Structural Insights on Retroviral DNA Integration: Learning from Foamy Viruses

Foamy viruses (FV) are retroviruses belonging to the Spumaretrovirinae subfamily. They are non-pathogenic viruses endemic in several mammalian hosts like non-human primates, felines, bovines, and equines. Retroviral DNA integration is a mandatory step and constitutes a prime target for antiretroviral therapy. This activity, conserved among retroviruses and long terminal repeat (LTR) retrotransposons, involves a viral nucleoprotein complex called intasome. In the last decade, a plethora of structural insights on retroviral DNA integration arose from the study of FV. Here, we review the biochemistry and the structural features of the FV integration apparatus and will also discuss the mechanism of action of strand transfer inhibitors

Activation and desensitisation of acetylcholine release by zinc at <i>Torpedo</i> nerve terminals

Treatment with 100 or 250 μM ZnCl2 irreversibly blocked neurotransmission in the Torpedo electric organ by inhibiting acetylcholine (ACh) release. In Zn2+-treated tissue, release failure did not result from impairment of Ca2+ entry since stimulation still provoked an accumulation of Ca2+. Also pretreatment of isolated synaptosomes with Zn2+ inhibited to the same extent the release elicited by KCl-evoked depolarisation and the release elicited by using the Ca2+ ionophore A23187. On the other hand, after application of A23187, Zn2+ by itself efficiently triggered ACh release from synaptosomes. This dual effect of Zn2+ was also observed to occur in proteoliposomes equipped with mediatophore (a protein of the presynaptic membrane characterised by its capability to support Ca2+-dependent transmitter release). Hence, Zn2+ mimicked two fundamental actions of Ca2+ on nerve terminals, which are: (1) the immediate activation of release, and (2) a more slowly developing desensitisation of release. Zn2+ was more powerful than Ca2+ for both actions. It is concluded that the dual action of Zn2+ on the mediatophore protein accounts at least in part for its complex effects on neurotransmission