Identification of human immunodeficiency virus type-1 Gag-TSG101 interaction inhibitors by high-throughput screening

Siarot, Lowela; Chutiwitoonchai, Nopporn; Sato, Hirotaka; Chang, Haiyan; Sato, Hironori; Fujino, Masayuki; Murakami, Tsutomu; Aono, Toshihiro; Kuroda, Kensuke; Takei, Masami; Aida, Yoko

doi:10.1016/j.bbrc.2018.08.079

Cited by 6 publications

(5 citation statements)

References 26 publications

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“…For instance, it has been proposed that targeting exosome biogenesis and release may have potential clinical implications for cancer therapy (26). The interaction between viral proteins and the ESCRT machinery has also been proposed as a potential target for antiviral therapy to fight against enveloped viruses (27,28) and also EV-associated naked viruses (29).…”

Section: Discussionmentioning

confidence: 99%

BK Polyomavirus Hijacks Extracellular Vesicles for En Bloc Transmission

Handala

Blanchard

Raynal

et al. 2020

J Virol

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Most people are asymptomatic carriers of the BK polyomavirus (BKPyV), but the mechanisms of persistence and immune evasion remain poorly understood. Furthermore, BKPyV is responsible for nephropathies in kidney transplant recipients. Unfortunately, the sole therapeutic option is to modulate immunosuppression, which increases the risk of transplant rejection. Using iodixanol density gradients, we observed that Vero and renal proximal tubular epithelial infected cells release two populations of infectious particles, one of which cosediments with extracellular vesicles (EVs). Electron microscopy confirmed that a single vesicle could traffic tens of viral particles. In contrast to naked virions, the EV-associated particles (eBKPyVs) were not able to agglutinate red blood cells and did not use cell surface sialylated glycans as an attachment factor, demonstrating that different entry pathways were involved for each type of infectious particle. However, we also observed that naked BKPyV and eBKPyV were equally sensitive to neutralization by the serum of a seropositive patient or commercially available polyvalent immunoglobulin preparations, which occurred at a postattachment step, after endocytosis. In conclusion, our work shows a new mechanism that likely plays a critical role during the primary infection and in the persistence, but also the reactivation, of BKPyV. IMPORTANCE Reactivation of BKPyV is responsible for nephropathies in kidney transplant recipients, which frequently lead to graft loss. The mechanisms of persistence and immune evasion used by this virus remain poorly understood, and a therapeutic option for transplant patients is still lacking. Here, we show that BKPyV can be released into EVs, enabling viral particles to infect cells using an alternative entry pathway. This provides a new view of BKPyV pathogenesis. Even though we did not find any decreased sensitivity to neutralizing antibodies when comparing EV-associated particles and naked virions, our study also raises important questions about developing prevention strategies based on the induction or administration of neutralizing antibodies. Deciphering this new release pathway could enable the identification of therapeutic targets to prevent BKPyV nephropathies. It could also lead to a better understanding of the pathophysiology of other polyomaviruses that are associated with human diseases.

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Section: Discussionmentioning

confidence: 99%

BK Polyomavirus Hijacks Extracellular Vesicles for En Bloc Transmission

Handala

Blanchard

Raynal

et al. 2020

J Virol

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“…We gave special attention to characterizing the interaction of Tsg101 with HIV-1 Gag in our system, because the essential role of Tsg101-Gag binding in HIV-1 replication makes it a potential target for new classes of antiretroviral therapy (27, 28). Similarly, because a number of other enveloped viruses ( e.g.…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, this cellular machinery is exploited by retroviruses such as HIV-1 and other enveloped viruses such as Ebola, which rely on the host ESCRT machinery to abscise virus particles from the surface of infected cells (21–25). Hijacking of the ESCRT machinery is critical to the replication of these viruses, and this host-pathogen interaction is not targeted by any of the current approved antiviral drugs, so it presents a potential target for new classes of antivirals (26–28). The many critical roles of the ESCRT machinery in human health and disease therefore make it imperative to understand the mechanisms of ESCRT function.…”

Section: Introductionmentioning

confidence: 99%

Genomic tagging of endogenous human ESCRT-I complex preserves ESCRT-mediated membrane-remodeling functions

Hoffman

Fernandez

Groves

et al. 2019

Journal of Biological Chemistry

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The endosomal sorting complexes required for transport (ESCRT) machinery drives membrane scission for diverse cellular functions that require budding away from the cytosol, including cell division and transmembrane receptor trafficking and degradation. The ESCRT machinery is also hijacked by retroviruses, such as HIV-1, to release virions from infected cells. The crucial roles of the ESCRTs in cellular physiology and viral disease make it imperative to understand the membrane scission mechanism. Current methodological limitations, namely artifacts caused by overexpression of ESCRT subunits, obstruct our understanding of the spatiotemporal organization of the endogenous human ESCRT machinery. Here, we used CRISPR/Cas9-mediated knock-in to tag the critical ESCRT-I component tumor susceptibility 101 (Tsg101) with GFP at its native locus in two widely used human cell types, HeLa epithelial cells and Jurkat T cells. We validated this approach by assessing the function of these knock-in cell lines in cytokinesis, receptor degradation, and virus budding. Using this probe, we measured the incorporation of endogenous Tsg101 in released HIV-1 particles, supporting the notion that the ESCRT machinery initiates virus abscission by scaffolding early-acting ESCRT-I within the head of the budding virus. We anticipate that these validated cell lines will be a valuable tool for interrogating dynamics of the native human ESCRT machinery.

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“…This protein-protein interaction thus constitutes an attractive drug target to develop antivirals. Peptidomimetics, including cyclic peptides (Tavassoli et al 2008;Lennard et al 2019) and small molecules (Siarot et al 2018), have indeed been shown to abolish the interaction between HIV-1 Gag and human Tsg101-UEV and to interfere with viral particles release. In addition, prazole-based drugs abolishing the ubiquitin binding function of Tsg101-UEV proved to interfere with the early HIV-1 assembly, independently of its interaction with the HIV-1 Gag PTAP late domain (Strickland et al 2017).…”

Section: Biological Contextmentioning

confidence: 99%

Backbone NMR resonance assignment of the apo human Tsg101-UEV domain

Moschidi¹,

Cantrelle²,

Boll³

et al. 2023

Biomol NMR Assign

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The Endosomal Sorting Complex Required for Transport (ESCRT) pathway, through inverse topology membrane remodeling, is involved in many biological functions, such as ubiquitinated membrane receptor trafficking and degradation, multivesicular bodies (MVB) formation and cytokinesis. Dysfunctions in ESCRT pathway have been associated to several human pathologies, such as cancers and neurodegenerative diseases. The ESCRT machinery is also hijacked by many enveloped viruses to bud away from the plasma membrane of infected cells. Human tumor susceptibility gene 101 (Tsg101) protein is an important ESCRT-I complex component. The structure of the Nterminal ubiquitin E2 variant (UEV) domain of Tsg101 (Tsg101-UEV) comprises an ubiquitin binding pocket next to a late domain [P(S/T)AP] binding groove. These two binding sites have been shown to be involved both in the physiological roles of ESCRT-I and in the release of the viral particles, and thus are attractive targets for antivirals. The structure of the Tsg101-UEV domain has been characterized, using X-ray crystallography or NMR spectroscopy, either in its apo-state or bound to ubiquitin or late domains. In this study, we report the backbone NMR resonance assignments, including the proline signals, of the apo human Tsg101-UEV domain, that so far was not publicly available. These data, that are in good agreement with the crystallographic structure of Tsg101-UEV domain, can therefore be used for further NMR studies, including protein-protein interaction studies and drug discovery.

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Identification of human immunodeficiency virus type-1 Gag-TSG101 interaction inhibitors by high-throughput screening

Cited by 6 publications

References 26 publications

BK Polyomavirus Hijacks Extracellular Vesicles for En Bloc Transmission

BK Polyomavirus Hijacks Extracellular Vesicles for En Bloc Transmission

Genomic tagging of endogenous human ESCRT-I complex preserves ESCRT-mediated membrane-remodeling functions

Backbone NMR resonance assignment of the apo human Tsg101-UEV domain

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