Projects - in details

Our studies focuse on the interferon signaling pathways/responses, and aims at understanding:

·       How do viruses escape or circumvent the host antiviral response within infected cells?

·       Discovery of the alternative pathways: how do cell types specialized for interferon production (e.g., plasmacytoid dendritic cells; pDCs) recognize the infected state of the neighboring cells?

We set up an unbiased experimental evolution in controlled cell environment to define the interactions between virus and host cells, and the routes of viral adaptation. Zika virus infection causes neurological diseases and birth defects, representing a threat for human health. The type I and III interferon responses are key for viral control, in utero transmission and Zika virus-induced neurological diseases. 

We conducted genomic screen-based studies of the Zika virus quasi-species evolution in response to a defined host environnement  to delineate the genetic interactions with host pathways in viral genome, and viral escape mechanisms. Bioinformatics analyses of deep-sequenced viral quasi-species pinpoint mutations that temporally coincided with acquisition of the adapted phenotype, idenitified as a faster initiation of replication associated to escape from antiviral responses. Mathematical modeling further guilded our  mutagenesis strategy using recombinant Zika virus molecular clone, leading to the functional validation of a key point mutation. Overall our multidsiciplinary approach provides insights into Zika virus adaptation to host cells and immune escape mechanisms, and can serve as a framework to study other viruses

Grass, et al. https://doi.org/10.1101/2020.12.28.424577

AIM II. SENSING OF INFECTED CELLS BY PLASMACYTOID DENDRITIC CELLS: CELL CONTACT-DEPENDENT AND ATYPICAL PAMP-CARRIERS

Virtually all viruses have evolved mechanisms to inhibit host-sensing pathways within cells they invade. Nonetheless, paradoxically, expressions of interferon and ISG are readily detected in infected patients. This suggests the existence of alternative sensing pathways.

Plasmacytoid dendritic cells (pDCs) are the more robust type I and III interferon (IFN-I/III) producers. Of note, since pDCs are refractory to virtually all viral infections, this sensing is unopposed by viral product, thus leading to robust antiviral response. Therefore, it opens questions : how pDCs sense immunostimulatory RNA (PAMPs) and prevent viral replication?

II-A. PAMP-CARRIERS: TRANSFER OF VIRAL IMMUNOSTIMULATORY SIGNAL VIA NON-CANONICAL AND/OR NON-INFECTIOUS VESICLES

Cells infected by Flavivirus release immature particles (i.e., with uncleaved viral membrane proteins, deficient for membrane fusion). This immature particles results from suboptimal cleavage site, which is evolutionarily conserved likely to prevent premature membrane fusion in the secretory pathway.

We demonstrated that immature particle is the PAMP-carrier, eliciting pDC response by the sensor TLR7, which is localized in the endolysosomes. This is in accordance with the fusion-deficiency of the immature virions, and thus their increased retention in the endolysosomal compartment and by consequence an augmented recognition of viral RNA by TLR7. Our finding might have broader importance for the many viruses that, similarly, prevent antiviral signaling within infected cells and release uncleaved glycoprotein-containing non-infectious particles.

Together with our previous studies, we thus revealed that the immunostimulatory signal can be transmitted via non-infectious and/or non-canonical carriers, i.e., immature virions and exosome in the context, respectively, of flavivirus and HCV, that we called PAMP-carriers.

Decembre, Assil et al. Plos Pathogens, 2014.

https://doi.org/10.1371/journal.ppat.1004434

Sinigaglia et al. Sci Rep. 2018.

https://doi.org/10.1038/s41598-018-29235-7

Dreux et al. Cell Host and Microbes. 2012.

http://doi.org/10.1016/j.chom.2012.08.010

Webster et al. J Virol. 2016.

http://doi.org/10.1128/JVI.01692-16

II-B. CELL SURFACE PATTERN OF GLYCOSYLATION regulate the SENSING OF CELL-ASSOCIATED HTLV BY PLASMACYTOID DENDRITIC CELLS  

Human T Leukemia virus type (HTLV) infection can persist, leading to adult T-cell Leukemia, a life-threatening cancer. Viral persistence results, at least in part, from viral escape of the innate immunity, including inhibition of IFN-I response in infected T-cells. We showed that pDCs sense surface associated-HTLV present within glycan-rich structure. Especially, the cell surface pattern of glycosylation modulates the transmission of PAMPs to pDCs. Thus, our results uncovered a function of virus-containing cell surface-associated glycosylated structures in the activation of innate immunity.

Assil et al. Plos Pathogens, 2019

https://doi.org/10.1371/journal.ppat.1004434

II.C. ANTIVIRAL RESPONSE BY PLASMACYTOID DENDRITIC CELLS VIA INTERFEROGENIC SYNAPSE WITH INFECTED CELLS.

While PAMP-carriers can be distinct e.g. immature virus & exosome (see above in II-A), physical contacts are required for the sensing by pDCs for genetically distant viruses. How and why these contacts are established ?

Using Dengue, hepatitis C (HCV) and Zika viruses as models, we demonstrated that the contact site of pDC with infected cell reorganizes into a specialized platform for the transfer of the PAMP-carrier, that we named interferogenic synapse. This newly defined synpase is required for pDC-induced antiviral response.

Assil, Coleon et al. Cell Host & Microbe 25, 730–745, 2019

https://doi.org/10.1016/j.chom.2019.03.005

We aim at defining the molecular basis of this newly discovered aspect of innate immunity.

AIM III. PLASMACYTOID DENDRITIC CELLS CONTROL ARBOVIRUSES VIA IRF7-INDUCED interferon RESPONSE

We demonstrated that sensing of infected cells by pDCs results in IRF7-driven IFN-I production in the absence of other inflammatory cytokine responses. To elucidate in vivo pDC immunomodulatory functions, we developed a mouse model, in which IRF7 signaling is restricted to pDCs. As viral model we studied Dengue and chikungunya viruses. Despite undetectable levels of type I interferon protein, pDC-restricted IRF7 signaling was sufficient to control both viruses and protect mice from lethal chikungunya infection. Early pDC IRF7-signaling resulted in amplification of downstream antiviral responses, including an accelerated NK cell-mediated type II interferon response. These studies revealed the dominant, yet indirect role of pDC IRF7-signaling in directing both type I and II interferon responses during arbovirus infections.

Webster et al eLIFE 2018

https://elifesciences.org/articles/34273

We now aim at understanding: how pDC IFN-I signaling locally mobilizes other host responses to control viral infection?

e.g., IFN-II by NK cells.

AIM IV. how TYPE I and III INTErFERON responseS control SarS-Cov2 infection and  severity of COVID-19 ?

This is a recent project, and the main objective is to define how the type I interferon response controls SARS-Cov2 and its pathogenesis at the molecular molecular level and combineb with a clinical study in a cohorte of COVID-19 patients. 

Sa Ribero et al Plos Pathogens 2020 

https://doi.org/10.1371/journal.ppat.1008737 

NEW AIM. Dynamics of the interferon response at the single cell level