Some aspects of the life of SARS-CoV-2 ORF3a protein in mammalian cells

Jiao, Song; Miranda, Pablo; Li, Yan; Maric, Dragan; Holmgren, Miguel

doi:10.1016/j.heliyon.2023.e18754

Cited by 5 publications

(4 citation statements)

References 73 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Again, the current density was not different from that recorded in control cells. The absence of ion currents when the 41 residues at the N-terminus were deleted, was consistent with a recent study showing that the same construct, which exhibited a robust cell surface expression, did not provide any additional currents compared to control cells 19 Thus, together with the structural analysis performed on the ORF3a protein (Fig. 1), our electrophysiological results strongly suggest that ORF3a protein is not an ion channel.…”

Section: Resultssupporting

confidence: 91%

“…Consistent with previous studies 6, 19 , the removal of this sequence substantially increased ORF3a localization on the plasma membrane, as clearly shown by the enhanced uorescence signal of cell surface exhibited by tORF3a-transfected cells (Fig. 2A, right), compared to ORF3a-transfected cells.…”

Section: Resultssupporting

confidence: 91%

“…Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is a positive-strand RNA β-coronavirus that causes coronavirus disease-19 (COVID- 19), responsible for the latest pandemic. Despite the numerous antiviral therapeutic strategies or vaccines under development, no fully effective anti-COVID-19 therapies have been developed to date.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

SARS-CoV-2 ORF3a accessory protein is a water-permeable channel that induces lysosome swelling

Michelucci,

Sforna,

Focaia

et al. 2024

Preprint

View full text Add to dashboard Cite

Open reading frame 3a (ORF3a) is among the most expressed viral accessory proteins of SARS-CoV-2, the pathogen responsible for the last pandemic. ORF3a mainly targets lysosomes of the host cell and promotes lysosomal inactivation through their deacidification, an essential step for lysosomal exocytosis and virus egress. However, the exact mechanism through which ORF3a performs this function is still unclear. While seminal studies suggested ORF3a functioning as a cation-selective viroporin, a recent work disproved this conclusion. To unravel the possible function of ORF3a, here we employed a multidisciplinary approach including molecular dynamics (MD), molecular biology, electrophysiology, and electron microscopy. Our electrophysiological results, in accordance with apreliminary MD structural analysis, ruled out that ORF3a functions as anion channel when expressed in HEK293 cells. Conversely, both MD and videoimaging experiments for the assessment of cell volume changes demonstrated that ORF3a mediates the transmembrane transport of water. Using MD, we also identified the putative selectivity filter for water permeation, and experimentally confirmed its relevance for water transport by showing that its mutation at the level of an asparagine (N82L and N82W) abolishes ORF3a-mediated water permeation. Finally, ORF3a expression in HEK293 cells determined lysosomal volume increase (swelling), mitochondrial damage, and accumulation of intracellular membranes, and these effects were reverted by the N82W mutation. Collectively, our data suggest a new function for the ORF3a protein as a lysosomal water permeable channel. The ORF3a-mediated water transport across lysosome membrane might promote lysosomal swelling and deacidification and, by consequence, inactivation, a key step to promote virus egress from the cell.

show abstract

Section: Resultssupporting

confidence: 91%

Section: Resultssupporting

confidence: 91%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

SARS-CoV-2 ORF3a accessory protein is a water-permeable channel that induces lysosome swelling

Michelucci,

Sforna,

Focaia

et al. 2024

Preprint

View full text Add to dashboard Cite

show abstract

“…We describe here a deep time-dependent proteome rewiring that drives a sequential reprogramming of cell biology to allow the progress of SARS-CoV-2 infection, which is in very good agreement with previous reports. Evasion of the cellular innate immunity, control of the translation machinery, regulation of the endocytic and secretory pathways, control of the apoptotic response and autophagy, induction ER stress and UPR, interference with the control of cell polarity and epithelial cell-cell junction integrity, lipid metabolism are the masterpieces of the host cell reprogramming by the virus, that is mediated by key viral proteins (22,(28)(29)(30)(31)(32)(33)(34)(35)(36). In addition to the expected accumulation of viral proteins over time, we also detected a time-dependent phosphorylation increase regulatory impact of which is unclear.…”

Section: Discussionmentioning

confidence: 99%

Time-based quantitative proteomic and phosphoproteomic analysis of A549-ACE2 cells during SARS-CoV-2 infection

Milhano dos Santos,

Vindel,

Ciordia

et al. 2024

Preprint

View full text Add to dashboard Cite

The outbreak of COVID-19, a disease caused by severe acute respiratory syndrome coronavirus 2, led to an ongoing pandemic with devastating consequences for the global economy and human health. With the global spread of SARS-CoV-2, multidisciplinary initiatives were launched to explore new diagnostic, therapeutic, and vaccination strategies. From this perspective, proteomics could help to understand the mechanisms associated with SARS-CoV-2 infection and to identify new therapeutic targets for antiviral drug repurposing and/or discovery. A TMT-based quantitative proteomics and phosphoproteomics analysis was performed to study the proteome remodeling of human lung alveolar cells transduced to express human ACE2 (A549-ACE2) after infection with SARS-CoV-2. Targeted PRM analysis was performed to assess the detectability in serum and prognostic value of selected proteins. A total of 6802 proteins and 6428 phospho-sites were identified in A549-ACE2 cells after infection with SARS-CoV-2. Regarding the viral proteome, 8 proteins were differentially expressed after 6 h of infection and reached a steady state after 9 h. In addition, we detected several phosphorylation sites of SARS-CoV-2 proteins, including two novel phosphorylation events at S410 and S416 of the viral nucleoprotein.ImportanceThe differential proteins here identified revealed that A549-ACE2 cells undergo a time-dependent regulation of essential processes, delineating the precise intervention of the cellular machinery by the viral proteins. From this mechanistic background and by applying machine learning modelling, 29 differential proteins were selected and detected in the serum of COVID-19 patients, 14 of which showed promising prognostic capacity. Targeting these proteins and the protein kinases responsible for the reported phosphorylation changes may provide efficient alternative strategies for the clinical management of COVID-19.

show abstract

Dynamic Cellular Proteome Remodeling during SARS-CoV-2 Infection. Identification of Plasma Protein Readouts

dos Santos,

Vindel-Alfageme,

Ciordia

et al. 2024

J. Proteome Res.

View full text Add to dashboard Cite

The outbreak of COVID-19, led to an ongoing pandemic with devastating consequences for the global economy and human health. With the global spread of SARS-CoV-2, multidisciplinary initiatives were launched to explore new diagnostic, therapeutic, and vaccination strategies. From this perspective, proteomics could help to understand the mechanisms associated with SARS-CoV-2 infection and to identify new therapeutic options. A TMT-based quantitative proteomics and phosphoproteomics analysis was performed to study the proteome remodeling of human lung alveolar cells expressing human ACE2 (A549-ACE2) after infection with SARS-CoV-2. Detectability and the prognostic value of selected proteins was analyzed by targeted PRM. A total of 6802 proteins and 6428 phospho-sites were identified in A549-ACE2 cells after infection with SARS-CoV-2. The differential proteins here identified revealed that A549-ACE2 cells undergo a time-dependent regulation of essential processes, delineating the precise intervention of the cellular machinery by the viral proteins. From this mechanistic background and by applying machine learning modeling, 29 differential proteins were selected and detected in the serum of COVID-19 patients, 14 of which showed promising prognostic capacity. Targeting these proteins and the protein kinases responsible for the reported phosphorylation changes may provide efficient alternative strategies for the clinical management of COVID-19.

show abstract

Some aspects of the life of SARS-CoV-2 ORF3a protein in mammalian cells

Cited by 5 publications

References 73 publications

SARS-CoV-2 ORF3a accessory protein is a water-permeable channel that induces lysosome swelling

SARS-CoV-2 ORF3a accessory protein is a water-permeable channel that induces lysosome swelling

Time-based quantitative proteomic and phosphoproteomic analysis of A549-ACE2 cells during SARS-CoV-2 infection

Dynamic Cellular Proteome Remodeling during SARS-CoV-2 Infection. Identification of Plasma Protein Readouts

Contact Info

Product

Resources

About