%0 Generic
%A Hoffmann, Anja
%D 2020
%F heidok:26778
%K Uukuniemi-Virus  Rab11  Atg7
%R 10.11588/heidok.00026778
%T Early phlebovirus host cell interactions:  The small GTPase Rab11 and the autophagic factor Atg7 promote Uukuniemi virus entry
%U https://archiv.ub.uni-heidelberg.de/volltextserver/26778/
%X Phleboviruses, in the Phenuiviridae family within the Bunyavirales order, are important pathogenic arthropod-borne viruses (arboviruses), causing severe diseases in humans and domestic animals. Outbreaks are no longer limited to tropical and developing countries. Global trade, deforestation and global warming are reasons for the expansion of arthropod vectors, and the viruses they carry. The mosquito-borne phlebovirus Rift Valley fever (RVFV) spread from sub-Saharan parts of Africa over the entire continent and to the Arabic peninsula during the last two decades. As it already happened for other arboviruses (e.g. dengue virus), RVFV is now at risk of introduction into Southern Europe. Phleboviruses represent a risk to public health and agricultural productivity and must be taken seriously as potential emerging and reemerging pathogens. For humans, neither specific antiviral treatments nor vaccines are currently approved.  Ideally, treating phlebovirus infection in humans, would target early virus-host cell interactions, preventing the release of the virus genome into the cytosol. Yet, the details of the entry pathways exploited by phleboviruses are mostly elusive, awaiting to be uncovered. For my PhD project, I used Uukuniemi virus (UUKV). UUKV is a validated biosafety level (BSL)-2 model for phleboviruses of higher biosafety classification such as RVFV.  Our lab previously reported that UUKV enters human host cells by receptor-mediated endocytosis, transits Rab5-positive early endosomes and penetrates the cytosol from late endosomal compartments with a pH value around 5.4. With the aim to identify additional host factors involved in UUKV entry, two genome-wide siRNA screens were performed. In those screens, VAMP3 was identified to facilitate late endosomal penetration of UUKV. The v-SNARE protein VAMP3 plays an important role in recycling endosome trafficking and the initiation of autophagy. In addition to VAMP3, several other autophagy-associated host factors were found as potential host factors in the siRNA screens for UUKV entry. The overall goal of my PhD project was to clarify the role of autophagy in phlebovirus entry and decipher the molecular mechanisms subverted by phleboviruses to penetrate human host cells. Therefore, I analyzed UUKV infection by flow cytometry and confocal microscopy approaches.  Within my PhD project, I assessed numerous autophagy-associated proteins for their role in UUKV infection. I identified the autophagic factor Atg7 and the small GTPase Rab11a as important host factors for UUKV infection. Atg7 is known mainly for its function in autophagosome maturation. Rab11a regulates recycling endosome trafficking and is involved in the initiation of autophagy. Addressing single steps of the virus entry process, I found that Atg7 and Rab11 specifically promote UUKV intracellular trafficking, while no effects were observed on other steps during early virus host cell interactions, i.e. binding or replication. Interestingly however, my results also indicate that Atg7 and Rab11 participate in UUKV infection in an autophagy-independent manner.  In conclusion, this thesis expands our knowledge about entry of UUKV particles into human cells with a role of two more host factors, Rab11a and Atg7. Both proteins facilitate the transport of endocytosed viral particles from the plasma membrane to acidic endosomal compartments. Reaching these compartments is a critical step for acid-activated fusion and the subsequent release of the viral genome into the cytosol. Additionally, this work provides an indication of autophagy-independent functions of Atg7 in endosomal trafficking. The importance of Rab11a and VAMP3 in UUKV infection points towards a potential involvement of recycling endosomes in UUKV intracellular trafficking. UUKV represents a tool of choice to better understand the role of recycling endosomes in late endosomal trafficking, a function that remains elusive and is potentially exploited by other related and unrelated viruses.