Biology subjects

Dancel-Manning, K.

Publications and source records attributed to Dancel-Manning, K..

3 recordsLinked to original sources

bioRxiv2021

ACE2-containing defensosomes serve as decoys to inhibit SARS-CoV-2 infection

Extracellular vesicles of endosomal origin, exosomes, mediate intercellular communication by transporting substrates with a variety of functions related to tissue homeostasis and disease. Their diagnostic and therapeutic potential has been recognized for diseases such as cancer in which signaling defects are prominent. However, it is unclear to what extent exosomes and their cargo inform the progression of infectious diseases. We recently defined a subset of exosomes termed defensosomes that are mobilized during bacterial infection in a manner dependent on autophagy proteins. Through incorporating protein receptors on their surface, defensosomes mediated host defense by binding and inhibiting pore-forming toxins secreted by bacterial pathogens. Given this capacity to serve as decoys that interfere with surface protein interactions, we investigated the role of defensosomes during infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of COVID-19. Consistent with a protective function, exosomes containing high levels of the viral receptor ACE2 in bronchioalveolar lavage fluid from critically ill COVID-19 patients was associated with reduced ICU and hospitalization times. We found ACE2+ exosomes were induced by SARS-CoV-2 infection and activation of viral sensors in cell culture, which required the autophagy protein ATG16L1, defining these as defensosomes. We further demonstrate that ACE2+ defensosomes directly bind and block viral entry. These findings suggest that defensosomes may contribute to the antiviral response against SARS-CoV-2 and expand our knowledge on the regulation and effects of extracellular vesicles during infection.

immunology↗

bioRxiv2021

Emerging chikungunya virus variants at the E1-E1 inter-glycoprotein spike interface impact virus attachment and Inflammation

Chikungunya virus (CHIKV) is a re-emerging arthropod-borne alphavirus and a serious threat to human health. Therefore, efforts toward elucidating how this virus causes disease and the molecular mechanisms underlying steps of the viral replication cycle are crucial. Using an in vivo transmission system that allows intra-host evolution, we identified an emerging CHIKV variant carrying a mutation in the E1 glycoprotein (V156A) in the serum of mice and saliva of mosquitoes. E1 V156A has since emerged in humans during an outbreak in Brazil, co-occurring with a second mutation, E1 K211T, suggesting an important role for these residues in CHIKV biology. Given the emergence of these variants, we hypothesized that they function to promote CHIKV infectivity and subsequent disease. Here, we show that E1 V156A and E1 K211T modulate virus attachment and fusion and impact binding to heparin, a homolog of heparan sulfate, a key entry factor on host cells. These variants also exhibit differential neutralization by anti-glycoprotein monoclonal antibodies, suggesting structural impacts on the particle that may be responsible for altered interactions at the host membrane. Finally, E1 V156A and E1 K211T exhibit increased titers in an adult arthritic mouse model and induce increased foot-swelling at the site of injection. Taken together, this work has revealed new roles for E1 where discrete regions of the glycoprotein are able to modulate cell attachment and swelling within the host. IMPORTANCEAlphaviruses represent a growing threat to human health worldwide. Chikungunya virus (CHIKV) has rapidly spread to new geographic regions in the last several decades, causing overwhelming outbreaks of disease, yet there are no approved therapeutics. The CHIKV glycoproteins are key determinants of CHIKV adaptation and virulence. In this study, we characterize the naturally emerging E1 glycoprotein variants, V156A and K211T. We demonstrate that E1 V156A and K211T function in virus attachment to cells, a role that until now has been only attributed to the CHIKV E2 glycoprotein. We also demonstrate E1 V156A and K211T to increase foot-swelling in mice. Observing that these variants and other pathogenic variants occur at the E1-E1 inter-spike interface, we highlight this structurally important region as critical for multiple steps during CHIKV infection. Together, these studies further defines the function of E1 in CHIKV infection and can inform the development of therapeutic or preventative strategies.

microbiology↗

bioRxiv2019

Human antiviral protein MxA forms novel metastable membrane-less cytoplasmic condensates exhibiting rapid reversible \"crowding\"-driven phase transitions

Phase-separated biomolecular condensates of proteins and nucleic acids form functional membrane-less organelles in the mammalian cell cytoplasm and nucleus. We report that the interferon (IFN)-inducible human \"myxovirus resistance protein A\" (MxA) forms membrane-less metastable condensates in the cytoplasm. Light and electron microscopy studies revealed that transient expression of HA- or GFP-tagged MxA in Huh7, HEK293T or Cos7 cells, or exposure of Huh7 cells to IFN-2a led to the appearance of MxA in the cytoplasm in membrane-less variably-sized spherical or irregular bodies, in filaments and even a reticulum. 1,6-Hexanediol treatment led to rapid disassembly of these condensates; however, FRAP revealed a relative rigidity with a mobile fraction of only 0.24{+/-}0.02 within condensates. In vesicular stomatitis virus (VSV)-infected Huh7 cells, the nucleocapsid (N) protein, which participates in forming phase-separated viral structures, associated with GFP-MxA condensates. Remarkably, the cytoplasmic GFP-MxA condensates disassembled within 1-3 min of exposure of cells to hypotonic medium (40-50 milliosmolar) and reassembled within 0.5-2 min of re-exposure of cells to isotonic medium (310-325 milliosmolar) through multiple cycles. Mechanistically, the extent of cytoplasmic \"crowding\" regulated this phase-separation process. GFP-MxA condensates also included the DNA sensor protein cyclic GMP-AMP synthase (cGAS), another protein known to be associated with liquid-like condensates. Functionally, GFP-MxA expression inhibited DNA/cGAS-responsive ISG54-luciferase activity but enhanced relative inducibility of ISG54-luc by IFN-, revealing a physical separation between condensate- and cytosol-based signaling pathways in the cytoplasm. Taken together, the data reveal a new aspect of the cell biology of MxA in the cell cytoplasm.\n\nImportanceThe human interferon-inducible \"myxovirus resistance protein A\" (MxA), which displays antiviral activity against several RNA and DNA viruses, exists in the cytoplasm in phase-separated membrane-less metastable condensates of variably-sized spherical or irregular bodies, in filaments and even in a reticulum. MxA condensate formation appeared necessary but not sufficient for antiviral activity. Remarkably, MxA condensates showed the unique property of rapid (within 1-3 min) reversible disassembly and reassembly in intact cells exposed sequentially to hypotonic and isotonic conditions Mechanistically, these phase transitions were regulated by the extent of cytoplasmic \"crowding.\" Moreover, GFP-MxA condensates included the DNA sensor protein cyclic GMP-AMP synthase (cGAS). Functionally, GFP-MxA expression inhibited DNA/cGAS-responsive ISG54-luciferase activity but enhanced inducibility of ISG54-luc by IFN-, revealing a biological distinction between condensate- and cytosol-based signaling pathways. Since intracellular edema and ionic changes are hallmarks of cytopathic viral effects, the rapid hypotonicity-driven disassembly of MxA condensates may modulate MxA.function during virus infection.

cell biology↗