Research interests

Our lab investigates the cell biology of arbovirus infections, using bunyaviruses as a model. The super group of arboviruses (for arthropod-borne viruses) comprises five families of viruses (Togaviridae, Rhabdoviridae, Reoviridae, Flaviviridae, and Bunyaviridae), with over a thousand members distributed worldwide. Some infamous members are West Nile, dengue, yellow fever, Chikungunya, Rift valley fever (RVFV), Schmallenberg, and Crimean–Congo hemorrhagic fever (CCHFV) viruses. These viruses are mainly transmitted by arthropods such as mosquitoes, sand-flies, and ticks. Many arboviruses cause severe illness in humans such as encephalitis, hepatitis, and hemorrhagic fever.

 

Due to their mode of transmission as well as global warming and human activity, these viruses are considered emerging agents of diseases. The flaviviruses dengue in Brazil and West Nile in USA (about 5,000 infected persons and 240 lethal cases in 2012) as well as the bunyaviruses Severe fever with thrombocytopenia syndrome (SFTSV) in China (2009), RVFV in South Africa (2010), and Schmallenberg in Europe (2011) are recent examples of worldwide outbreaks demonstrating that arboviruses must be taken seriously as potential emerging agents of disease. Currently there are no approved vaccines or treatments to protect humans from infection.

As for many other arboviruses, research on bunyaviruses has not had a high priority in the past, and therefore relatively little is known about their strategies for transmission and infection.

 

Bunyaviridae

 

Bunyaviruses are enveloped and spherical (about 100 nm in diameter), with a segmented negative-sense RNA genome. The three RNA segments replicate in the cytosol and encode four structural proteins, the nucleoprotein N, the RNA-dependent RNA polymerase, and two transmembrane glycoproteins (Gn and Gc). In the envelope, the glycoproteins form spike-like projections responsible for virus attachment to target cells and acid-activated membrane fusion. Based on computational modeling and the X-ray structure of RVFV glycoprotein C, it is believed that bunyavirus glycoproteins Gc are class II fusion proteins.

 

Electron micrograph of the phlebovirus Uukuniemi, a model for RVFV and SFTSV, two important human pathogens.

 

 

 

 

 

 

 

 

 

 

 

 

 

During natural transmission, arboviruses are introduced into the skin, following the bite of infected arthropods. Due to their presence in the anatomical site of initial infection, skin dermal dendritic cells (DCs) are among the first host cells to encounter the viruses and interactions between DCs and arboviruses are thought to be particularly relevant. Alpha-, bunya-, and flaviviruses have, in fact, been shown to infect immature DCs.

 

Bunyavirus entry

 

The receptors, cellular factors, and pathways used by bunyaviruses to enter their host cells remain largely unidentified and poorly characterized. However, many bunyaviruses use the C-type lectin CD209 as a receptor to enter DCs and some members of the hantavirus subfamily have been shown to use the 3 integrin to infect endothelial cells. Heparan sulfate has been implicated in RVFV attachment to cells. Sensitivity to perturbation of clathrin and dynamin-2 has implied a role for clathrin-mediated endocytosis in uptake and internalization.

Several lines of evidence suggest that many bunyaviruses are late-penetrating viruses (L-PVs), a large group of viruses that depends on trafficking to late endosomes for productive infection. Inhibitor studies have shown that infection relies on functional microtubules and vacuolar acidification. Weak bases that neutralize endosomal pH, such as ammonium chloride (NH4Cl), inhibit infection by most bunyaviruses. Acid-activated viral membrane fusion that results in the release of the virus genome in the cytosol occurs from late endosomal vacuole at pH values typically below 5.8.

 

Uukuniemi virus (red) in LAMP-1+ late endosomal vesicles (green).

 

 

 

 

 

 

 

 

 

 

 

Research program

We have interest in all steps of early arbovirus-host interactions, from the virus transmission to the genome release into the cytosol. Using cellular and molecular techniques in combination with video microscopy in live cells, we investigate how virus particles bind to cells, how they are internalized by endocytosis, and how they are transported to various organelles such as endosomes. We are also analyzing how the viral genome and accessory proteins escape into the cytosol. Through this research program, we expect to gain a detailed picture of the molecular and cellular mechanisms subverted by these viruses to infect a host.

Our current work is focused on the phleboviruses Uukuniemi, Punta Toro, and Toscana as well as the nairovirus Hazara. We are also interested in the phlebovirus Rift valley fever and the flavivirus West Nile.

 

 

Early host-arbovirus interactions, from transmission to genome release.