Alphavirus-Induced Membrane Rearrangements during Replication, Assembly, and Budding

Elmasri, Zeinab; Nasal, Benjamin L; Jose, Joyce

doi:10.3390/pathogens10080984

Cited by 12 publications

(10 citation statements)

References 189 publications

(273 reference statements)

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“…In mosquito cells, virus budding occurs at the plasma membrane as well as at the membranes of large cytoplasmic vesicles [ 15 ]. Electron microscopy (EM) analyses of alphavirus-infected mammalian cells have revealed two types of virus-induced structures called cytopathic vacuoles type 1 (CPV-Is) and type II (CPV-IIs) [ 16 ]. CPV-Is are larger and associated with replication complexes (RC) [ 15 , 17 , 18 ].…”

Section: Introductionmentioning

confidence: 99%

Requirement of a functional ion channel for Sindbis virus glycoprotein transport, CPV-II formation, and efficient virus budding

et al. 2022

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Many viruses encode ion channel proteins that oligomerize to form hydrophilic pores in membranes of virus-infected cells and the viral membrane in some enveloped viruses. Alphavirus 6K, human immunodeficiency virus type 1 Vpu (HIV-Vpu), influenza A virus M2 (IAV-M2), and hepatitis C virus P7 (HCV-P7) are transmembrane ion channel proteins that play essential roles in virus assembly, budding, and entry. While the oligomeric structures and mechanisms of ion channel activity are well-established for M2 and P7, these remain unknown for 6K. Here we investigated the functional role of the ion channel activity of 6K in alphavirus assembly by utilizing a series of Sindbis virus (SINV) ion channel chimeras expressing the ion channel helix from Vpu or M2 or substituting the entire 6K protein with full-length P7, in cis. We demonstrate that the Vpu helix efficiently complements 6K, whereas M2 and P7 are less efficient. Our results indicate that while SINV is primarily insensitive to the M2 ion channel inhibitor amantadine, the Vpu inhibitor 5-N, N-Hexamethylene amiloride (HMA), significantly reduces SINV release, suggesting that the ion channel activity of 6K is similar to Vpu, promotes virus budding. Using live-cell imaging of SINV with a miniSOG-tagged 6K and mCherry-tagged E2, we further demonstrate that 6K and E2 colocalize with the Golgi apparatus in the secretory pathway. To contextualize the localization of 6K in the Golgi, we analyzed cells infected with SINV and SINV-ion channel chimeras using transmission electron microscopy. Our results provide evidence for the first time for the functional role of 6K in type II cytopathic vacuoles (CPV-II) formation. We demonstrate that in the absence of 6K, CPV-II, which originates from the Golgi apparatus, is not detected in infected cells, with a concomitant reduction in the glycoprotein transport to the plasma membrane. Substituting a functional ion channel, M2 or Vpu localizing to Golgi, restores CPV-II production, whereas P7, retained in the ER, is inadequate to induce CPV-II formation. Altogether our results indicate that ion channel activity of 6K is required for the formation of CPV-II from the Golgi apparatus, promoting glycoprotein spike transport to the plasma membrane and efficient virus budding.

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Section: Introductionmentioning

confidence: 99%

Requirement of a functional ion channel for Sindbis virus glycoprotein transport, CPV-II formation, and efficient virus budding

et al. 2022

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“…We therefore conclude that BNIP3 possibly interferes with the stages occurring after the nucleocapsid is delivered into the cytoplasm and prior to robust RNA replication and translation. Early alphavirus RNA synthesis starts in spherules located on the plasma membrane, which are then internalized in an actin-myosin dependent manner and form cytopathic vacuoles, where RNA replication continues [76, 77]. Therefore, an intact actin network is found to be essential for the trafficking of alphavirus replication complexes [78].…”

Section: Discussionmentioning

confidence: 99%

BNIP3 regulates Chikungunya virus infection independently of autophagy and cell death

Echavarría-Consuegra

Kumar

Mauthe

et al. 2022

Preprint

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Chikungunya virus (CHIKV) is a human pathogen causing outbreaks of febrile illness for which vaccines and specific treatments remain unavailable. Autophagy-related (ATG) proteins and autophagy receptors are a set of host factors that participate in autophagy, but have also shown to function in other unrelated cellular pathways. Although autophagy is reported to both inhibit and enhance CHIKV replication, the specific role of individual ATG proteins remains largely unknown. Here, a siRNA screen was performed to evaluate the importance of the ATG proteome and autophagy receptors in controlling CHIKV infection. We observed that 7 out of 50 ATG proteins impact the replication of CHIKV. Among those, depletion of the mitochondrial protein and autophagy receptor BCL2 Interacting Protein 3 (BNIP3) increased CHIKV infection. Interestingly, BNIP3 controls CHIKV independently of autophagy and cell death. Detailed analysis of the CHIKV viral cycle revealed that BNIP3 interferes with the early stages of infection. Moreover, the antiviral role of BNIP3 was found conserved across two distinct CHIKV genotypes and the closely related Semliki Forest virus. Altogether, this study describes a novel and previously unknown function of the mitochondrial protein BNIP3 in the control of the early stages of the alphavirus viral cycle.

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“…Major differences exist between alphavirus-induced mammalian and mosquito membrane rearrangements, so experimental characterization is required to understand the relevance of these cleavages in each host. [139, 140]…”

Section: Discussionmentioning

confidence: 99%

Alphavirus nsP2 protease structure and cleavage prediction: Possible relevance to the pathogenesis of viral arthritis

Prescott¹

2022

Preprint

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Alphaviruses are a diverse genus of arboviruses capable of infecting many vertebrates including humans. Human infection is common in equatorial and subtropical regions and is often accompanied by arthralgia or encephalitis depending on viral lineage. No antivirals or vaccines have been approved, and many alphavirus lineages have only recently been discovered and classified. Alphavirus nsP2 protease is an important virulence factor yet is commonly thought to be a simple papain-like protease which only cleaves viral polyproteins. Here, I reveal novel molecular mechanisms of these proteases via sequence and predicted structure alignment and propose novel cellular mechanisms for the pathogenesis of viral arthritis by predicting which human proteins are likely cleaved by these proteases. In addition to the known primary cysteine mechanism in all alphaviruses and a secondary serine mechanism documented in chikungunya virus (CHIKV), I discovered secondary cysteine and threonine mechanisms exist in many other alphaviruses and that these secondary mechanisms coevolve with their viral polyprotein cleavages. As for cleavage prediction, neural networks trained on 93 different putative viral polyprotein cleavages achieved a Matthews correlation coefficient of 0.965, and, when applied to the human proteome, predicted that hundreds of proteins may be vulnerable. Notable pathways likely affected by cleavages include the cytoskeleton and extracellular matrix, antiproteases, protein translation/folding/glycosylation/ubiquitination, cellular differentiation, inflammation, and vesicle trafficking, hinting that this viral protease is a more important virulence factor than previously believed.

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Alphavirus-Induced Membrane Rearrangements during Replication, Assembly, and Budding

Cited by 12 publications

References 189 publications

Requirement of a functional ion channel for Sindbis virus glycoprotein transport, CPV-II formation, and efficient virus budding

Requirement of a functional ion channel for Sindbis virus glycoprotein transport, CPV-II formation, and efficient virus budding

BNIP3 regulates Chikungunya virus infection independently of autophagy and cell death

Alphavirus nsP2 protease structure and cleavage prediction: Possible relevance to the pathogenesis of viral arthritis

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