Proteomic approaches to uncovering virus–host protein interactions during the progression of viral infection

May combine two claims.

AP-MS can be used either to study interactions during infection, providing temporal information, or after overexpression of individual viral gene products to study single protein functions.

“This approach has been used to either study interactions during the progression of an infection, providing information about the temporality of interaction events, or following the overexpression of individual viral gene products to gain insight into the functions of single proteins (Figure 2A).”

May combine two claims.

AP-MS has been successfully used in virology to isolate virus-virus and virus-host multi-protein complexes, enabling identification of both indirect and direct protein interactions.

“AP-MS has been successfully implemented in virology studies to isolate virus–virus and virus–host multi-protein complexes, allowing the identification of both indirect and direct protein interactions.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Proteomic approaches have been increasingly applied to virology studies over the last decade.

“The last decade has witnessed the increasing application of proteomic approaches to virology studies [22].”

• cites [22] sourceNot checked yet

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Probably not a substantial claim: may not pass S4 (based). May combine two claims.

All viruses are obligate parasites whose evolutionarily conserved life cycles depend on virus-host interactions, often through protein-protein associations.

“In fact, all viruses are obligate parasites and undergo evolutionarily conserved life cycles that dutifully depend on virus–host interactions, frequently mediated by protein–protein associations.”

May combine two claims.

Temporally and spatially regulated virus-host protein interactions are central to virus replication and spread and to modulation of host antiviral defenses.

“At the core of virus replication and spread is the formation of virus–host protein interactions.”

• Also stated in: § Introduction · plain

  “Therefore, the establishment of virus–host and host–host protein interactions that are temporally and spatially regulated is critical for the progression of the virus life cycle, as well as for the modulation of host antiviral defenses.”

May combine two claims.

In PRV-infected neurons, Us9 interacts with KIF1A, and KIF1A is required for viral capsid transport in axons and for infection spread.

“the Enquist research group found that the viral protein Us9 interacts with the cellular kinesin KIF1A.”

• Also stated in: § Identifying protein interactions using epitope-tagged viruses · boosted

  “demonstrating that KIF1A is required for the transport of viral capsid in axons and for the spread of infection [42].”

• cites [42] sourceNot checked yet

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May combine two claims.

In HCMV-infected primary human fibroblasts, the HDAC1-containing nucleosome remodeling and deacetylase complex interacts with viral proteins pUL29/28 and pUL38, an association implicated in facilitating immediate-early viral RNA production.

“By AP-MS, it was observed that the HDAC1-containing nucleosome remodeling and deacetylase complex interacted with viral proteins pUL29/28 and pUL38. This association was ultimately implicated in facilitating the production of immediate-early viral RNAs.”

• cites [38] Terhune SS, 2010, Plos Pathog sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Magnetic beads are versatile for capturing multi-protein complexes of various sizes and reduce nonspecific associations through washing.

“By providing surface binding, these beads tend to be versatile for capturing multi-protein complexes of various sizes. Additionally, given their isolation via a magnet, these beads can be readily subjected to washing steps, reducing the presence of nonspecific associations [23].”

• cites [23] sourceNot checked yet

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May combine two claims.

In SINV-infected fibroblasts, nsP3 interacts with G3BP at all assessed time points (2-10 hpi) and with 14-3-3 only at late 6, 8, and 10 hpi.

“This study led to the discovery of time-specific interactions, with the early and stable recruitment of an endoribonuclease (G3BP), identified at all time points assessed, and the later recruitment of signaling proteins (14-3-3), identified only at the late 6, 8, and 10 hpi time points in the SINV life cycle.”

• cites [24] sourceNot checked yet

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May combine two claims.

Yeast two-hybrid and cross-linking are proteomic methods for identifying direct protein interactions during infection.

“Such methods include yeast-two hybrid (Y2H) and cross-linking (Figure 2B)”

May combine two claims.

In HCMV-infected primary human fibroblasts, pUL27 is necessary and sufficient to destabilize and degrade cellular acetyltransferase Tip60.

“pUL27, a protein of previously unknown function, was found to be necessary and sufficient to destabilize and degrade a cellular acetyltransferase, Tip60.”

• cites [39] sourceNot checked yet

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May combine two claims.

In HSV-1-infected human epidermoid cells at 6 hpi, ICP27 interacts with eIF3 subunits p47 and p116, eIF4G, and poly-adenylate binding protein, suggesting ICP27 may interfere with viral or host mRNA translation.

“the Knipe group subsequently identified the eukaryotic translation initiation eIF3 subunits p47 and p116, eIF4G, and poly-adenylate binding protein as cellular interactors of the HSV-1 immediate-early protein ICP27 in human epidermoid cells at 6 hpi. These results suggested that ICP27 may interfere with viral or host mRNA translation [36].”

• cites [36] sourceNot checked yet

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May combine two claims.

In HSV-1-infected human fibroblasts, ICP0 targets viral protein pUL46 for proteasome-dependent degradation, proposed as internal regulation of viral protein levels.

“the viral E3 ubiquitin ligase protein ICP0 does not only target host defense proteins for proteasome-dependent degradation, but also a viral protein, the outer tegument protein pUL46.”

• Also stated in: § Identifying protein interactions using epitope-tagged viruses · hedged

  “The interaction between pUL46 and ICP0, observed and validated at both time points, was further functionally characterized and proposed to likely act as an internal regulation of viral protein levels during the virus life cycle [40].”

• cites [40] sourceNot checked yet

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May combine two claims.

Cryogenic cell lysis provides reproducible and effective cell disruption, helps access challenging protein interactions, and preserves protein complexes and weak interactions.

“this strategy was shown to help preserve protein complexes and weak protein interactions [24,25].”

• cites [24] sourceNot checked yet

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• cites [25] sourceNot checked yet

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May combine two claims.

In HSV-1-infected cells, ICP0 is the most abundant viral interaction with IFI16 at 3 and 8 hpi, and ICP0 targets IFI16 for proteasome-dependent degradation, inhibiting IFI16-mediated immune responses.

“the HSV-1 immediate-early transactivating protein ICP0 was determined to be the most abundant viral interaction with IFI16. Further functional analyses [18] provided additional support for the current model that during early stages of HSV-1 infection, ICP0 employs a virus immune evasion strategy mediated by protein associations to target IFI16 for proteasome-dependent degradation. This early assoc”

• cites [18] sourceNot checked yet

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• cites [19] sourceNot checked yet

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• cites [47] sourceNot checked yet

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May combine two claims.

In HCV-infected hepatoma cells, CD81 interacts with serum response factor binding protein 1, which is critical for HCV cell penetration and is a specific entry factor for all seven HCV genotypes.

“Serum response factor binding protein 1 was identified as a CD81 interaction, and shown to have a critical role in the cell penetration by HCV. This protein was further demonstrated to be a specific virus entry factor for HCV, functioning for all seven HCV genotypes.”

• cites [32] sourceNot checked yet

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May combine two claims.

In cells overexpressing BEFVα1, BEFVα1 interacts with importins β1 and 7, implicating nuclear trafficking in infection progression.

“GFP tagging and AP-MS led to the identification of a novel association between a protein of unknown function in the rhabdovirus bovine ephemeral fever virus, BEFVα1, and the cellular importins β1 and 7, implicating nuclear trafficking in the progression of infection [41].”

• cites [41] sourceNot checked yet

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May combine two claims.

Overexpression of single epitope-tagged viral proteins is an alternative when full-length replication-competent virus strains are unavailable, but it has a different biological impact than infection, limiting data interpretation and potentially yielding biologically irrelevant and false-negative associations.

“It is important to consider that this approach has a different biological impact on the host cell than an infection with a full-length virus, thereby limiting the data interpretation.”

• Also stated in: § AP-MS protein interaction studies using overexpression of individual viral gene products · hedged

  “both biologically irrelevant and false-negative associations may arise in a context-dependent manner.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Sequestration of importins in the cytoplasm has been reported for Ebola as a means to inhibit interferon signaling.

“The sequestration of importins in the cytoplasm has been reported for other non-nuclear-replicating viruses, such as Ebola [58], as a means to inhibit interferon signaling.”

• cites [58] sourceNot checked yet

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May combine two claims.

Overexpression of BEFVα1 slightly decreases nuclear importin β1 levels but does not affect importin 7 localization.

“showing that overexpression of BEFVα1 leads to a slight decrease in the levels of nuclear importin β1, while not affecting the localization of importin 7.”

• cites [41] sourceNot checked yet

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May combine two claims.

In HCMV infection, the major viral tegument protein binds IFI16 in the nucleus, blocking its oligomerization and antiviral cytokine response.

“the major viral tegument protein during HCMV infection was shown to bind to IFI16 in the nucleus, thereby blocking its ability to oligomerize and initiate antiviral cytokine response.”

• cites [48] sourceNot checked yet

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• cites [49] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The glycoprotein spike proteins of other coronaviruses associate with cellular receptors, facilitating virion entry.

“It was previously known that the glycoprotein spike proteins of other coronaviruses associate with cellular receptors, facilitating virion entry [60–63].”

• cites [60] sourceNot checked yet

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• cites [61] sourceNot checked yet

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• cites [62] sourceNot checked yet

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• cites [63] Kubo H, 1994, J Virol sourceNot checked yet

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May combine two claims.

By assessing various HPV subtypes, these studies were able to identify both strain-specific associations, as well as interactions conserved across HPV strains.

“By assessing various HPV subtypes, clinically classified by severity of oncogenesis and disease, these studies were able to identify both strain-specific associations, as well as interactions conserved across HPV strains.”

• cites [64] sourceNot checked yet

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• cites [65] sourceNot checked yet

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May combine two claims.

Subsequent functional assays further suggested that the virus may sequester USP7 indirectly to inhibit host-induced apoptosis and promote cell cycle growth.

“Subsequent functional assays further suggested that the virus may sequester USP7 indirectly to inhibit host-induced apoptosis and promote cell cycle growth.”

• cites [71] sourceNot checked yet

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May combine two claims.

TAP tagging can help in obtaining cleaner purifications in comparison to one-step isolations and for identifying strong protein-protein interactions.

“this method can help in obtaining cleaner purifications in comparison to one-step isolations and for identifying strong protein–protein interactions.”

Probably not a substantial claim: may not pass S3 (load-bearing).

TAP tagging was first used to identify multiprotein yeast complexes in 1999.

“TAP tagging was first used to identify multiprotein yeast complexes in 1999 [70]”

• cites [70] sourceNot checked yet

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May combine two claims.

As SAMHD1 hinders efficient viral cDNA synthesis during HIV infection, this study illustrates a viral usurpation mechanism by which HIV-2 evolved to establish infection in myeloid cells by inhibiting the host innate immune response.

“As SAMHD1 hinders efficient viral cDNA synthesis during HIV infection, this study illustrates a viral usurpation mechanism by which HIV-2 evolved to establish infection in myeloid cells by inhibiting the host innate immune response.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

While some viral proteins can be tagged at their C- or N-termini, others may require the inclusion of the tag within an internal loop.

“While some viral proteins can be tagged at their C- or N-termini (e.g. [86]), others may require the inclusion of the tag within an internal loop (e.g. [24]).”

• cites [86] sourceNot checked yet

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• cites [24] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

These findings corroborated previous observations that VP40 is sufficient to generate Ebola virus-like particles that form by budding off from host plasma membranes.

“These findings corroborated previous observations that VP40 is sufficient to generate Ebola virus-like particles that form by budding off from host plasma membranes [79,80].”

• cites [79] sourceNot checked yet

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• cites [80] sourceNot checked yet

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Classified as probably not a substantial claim.

It was previously reported that in myeloid cells transduced with HIV-2, Vpx prevents the inhibition of HIV-1 infection by associating with and hijacking a cellular E3 ubiquitin-protein ligase complex, including the proteins cullin 4 and DCAF1.

“It was previously reported that in myeloid cells transduced with HIV-2, Vpx prevents the inhibition of HIV-1 infection by associating with and hijacking a cellular E3 ubiquitin-protein ligase complex, including the proteins cullin 4 and DCAF1 [82,83].”

• cites [82] sourceNot checked yet

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• cites [83] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The use of a smaller tag tends to be preferred, as a large tag may impact protein size, affecting its localization and interactions.

“However, the use of a smaller tag tends to be preferred, as a large tag may impact protein size, affecting its localization and interactions.”

Probably not a substantial claim: may not pass S2 (checkable).

The global strategy adopted by this study and others provided a resource for further hypothesis-driven investigations into the functions of both characterized and poorly understood proteins during viral infection.

“The global strategy adopted by this study and others provided a resource for further hypothesis-driven investigations into the functions of both characterized and poorly understood proteins during viral infection.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Of these interactions, six were in complexes involved in DNA replication and packaging of phage particles.

“Of these interactions, six were in complexes involved in DNA replication and packaging of phage particles.”

• cites [87] sourceNot checked yet

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May combine two claims.

The available location for tagging is influenced by the protein conformation and by the necessity of certain domains for protein interactions.

“The available location for tagging is influenced by the protein conformation and by the necessity of certain domains for protein interactions.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Even the inability to tag a protein at a given location can be informative and highlight a functionally important region.

“Therefore, even the inability to tag a protein at a given location can be informative and highlight a functionally important region.”

May combine two claims.

NS3 and NS5A were found to deregulate cellular focal adhesion, which could instigate tumorigenesis and cell detachment from extracellular matrix components.

“Interestingly, NS3 and NS5A were found to deregulate cellular focal adhesion, which could instigate tumorigenesis and cell detachment from extracellular matrix components [89].”

• cites [89] sourceNot checked yet

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May combine two claims.

AP-MS approaches can reveal the formation of virus-host macromolecular complexes during infection, and inform on downstream pathways regulated by certain functional protein complexes.

“The use of AP-MS approaches can reveal the formation of virus–host macromolecular complexes during infection, and inform on downstream pathways regulated by certain functional protein complexes.”

Probably not a substantial claim: may not pass S3 (load-bearing).

A larger tag tends to offer higher efficiency of isolation, as the use of polyclonal antibodies or multiple monoclonal antibodies is possible.

“A larger tag tends to offer higher efficiency of isolation, as the use of polyclonal antibodies or multiple monoclonal antibodies is possible.”

Probably not a substantial claim: may not pass S3 (load-bearing).

As early as 1996, Bartel et al. applied a genome-wide Y2H screen on proteins from E. coli and bacteriophage T7 to reveal 25 interactions.

“As early as 1996, Bartel et al. applied a genome-wide Y2H screen on proteins from Escherichia coli and bacteriophage T7 to reveal 25 interactions [87].”

• cites [87] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The isolation of protein complexes by AP-MS is known to be affected by the generation of non-specific associations during cell lysis.

“the isolation of protein complexes by AP-MS is known to be affected by the generation of non-specific associations during cell lysis, as detailed above.”

May combine two claims.

The integration of AP-MS with Y2H can help to filter some of the false positives and nonspecific associations generated by these approaches, and highlight the most prominent interactions for further functional analyses.

“The integration of AP-MS with Y2H can help to filter some of the false positives and nonspecific associations generated by these approaches, and highlight the most prominent interactions for further functional analyses.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Protein cross-linking has been instrumental in uncovering numerous protein-protein interactions.

“Beginning as early as the 1960s, protein cross-linking has been instrumental in uncovering numerous protein–protein interactions [95,96].”

• cites [95] sourceNot checked yet

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• cites [96] sourceNot checked yet

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May combine two claims.

From this study, researchers identified over 1600 virus-host protein associations that highlighted both species-specific and conserved viral immune-evasive strategies.

“From this study, researchers identified over 1600 virus–host protein associations that highlighted both species-specific and conserved viral immune-evasive strategies [92].”

• cites [92] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

Formaldehyde crosslinking has become an integral element in numerous fundamental biochemical, molecular, and cellular biological techniques.

“As an example, formaldehyde crosslinking has become an integral element in numerous fundamental biochemical, molecular, and cellular biological techniques”

• cites [98] sourceNot checked yet

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May combine two claims.

This method covalently links one protein to another in virus-infected cells, thereby preserving weak or dynamic noncovalent protein interactions.

“This method covalently links one protein to another in virus-infected cells, thereby preserving weak or dynamic noncovalent protein interactions.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Y2H is known to generate false positives as proteins that may not be present in the same subcellular compartment can be artificially made to associate.

“Y2H is known to generate false positives as proteins that may not be present in the same subcellular compartment can be artificially made to associate.”

May combine two claims.

Cross-linking has been effective in capturing transient or weak intermolecular protein complexes, and gaining knowledge of intramolecular surface topologies, structural conformations, as well as of the interacting amino acid residues.

“This approach has been effective in capturing transient or weak intermolecular protein complexes, and gaining knowledge of intramolecular surface topologies, structural conformations, as well as of the interacting amino acid residues [97].”

• cites [97] sourceNot checked yet

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May combine two claims.

By these means, the authors cross-linked proteins in a purified potato leafroll virus sample and revealed topological interaction data in the virus that are required for virus-plant associations and its transmission across aphid vectors.

“By these means, the authors cross-linked proteins in a purified potato leafroll virus sample and revealed topological interaction data in the virus that are required for virus–plant associations and its transmission across aphid vectors [100].”

• cites [100] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The identification of an association does not inform if this interaction is direct or indirect.

“the identification of an association does not inform if this interaction is direct or indirect”

May combine two claims.

NAPPA arrays containing 10,000 purified human cDNA plasmids were incubated with rubella virus-infected cell lysates, leading to the identification and then confirmation of interactions between the viral capsid protein and host proteins.

“For example, NAPPA arrays containing 10,000 purified human cDNA plasmids were incubated with rubella virus-infected cell lysates, leading to the identification and then confirmation of interactions between the viral capsid protein and host proteins [104].”

• cites [104] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The comparison of the proteins identified in the bait and control isolations can be performed using label-free approaches, such as comparing spectral counts or precursor ion intensities obtained from LC-MS analyses.

“The comparison of the proteins identified in the bait and control isolations can be performed using label-free approaches, such as comparing spectral counts or precursor ion intensities obtained from LC–MS analyses [28].”

• cites [28] sourceNot checked yet

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May combine two claims.

These comparative analyses can help remove likely non-specific associations and uncover the protein interactions that are specifically enriched with the bait viral or host protein of interest.

“These comparative analyses can help remove likely non-specific associations and uncover the protein interactions that are specifically enriched with the bait viral or host protein of interest.”

Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing).

As this resource is still growing and the available number of controls for infected cells is limited, caution has to be taken when using this repository for predicting nonspecific associations during viral infection.

“However, as this resource is still growing and the available number of controls for infected cells is limited, caution has to be taken when using this repository for predicting nonspecific associations during viral infection.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Control AP-MS isolations are performed in parallel to the isolation of the viral or host protein of interest.

“For example, control AP-MS isolations are performed in parallel to the isolation of the viral or host protein of interest.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Several research groups have put together a resource of AP-MS controls, termed the contaminant repository for affinity purification.

“In an attempt to predict the associations that are likely to be non-specific, several research groups have put together a resource of AP-MS controls, termed the contaminant repository for affinity purification [27].”

• cites [27] sourceNot checked yet

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Probably not a substantial claim: may not pass S2 (checkable).

Once the subset of predicted specific interactions have been identified, it is imperative to confirm the associations of interest by orthogonal experimental methods.

“Once the subset of predicted specific interactions have been identified, it is imperative to confirm the associations of interest by orthogonal experimental methods.”

Probably not a substantial claim: may not pass S3 (load-bearing).

This resource provides useful information about recurring sticky proteins in different cell types or when using certain tags or resins for the isolations.

“This resource provides useful information about recurring sticky proteins in different cell types or when using certain tags or resins for the isolations.”

• cites [27] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

A more precise comparison of the bait and control IPs can be provided by relative quantification using metabolic labeling with stable isotopes.

“Alternatively, a more precise comparison of the bait and control IPs can be provided by relative quantification using metabolic labeling with stable isotopes [31,32]”

• cites [31] sourceNot checked yet

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• cites [32] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

The identification of protein-protein interactions using either AP-MS, Y2H, cross-linking or protein array studies can provide critical biological insight into protein function.

“The identification of protein–protein interactions using either AP-MS, Y2H, cross-linking or protein array studies can provide critical biological insight into protein function.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Interactions dependent on posttranslational modifications may not always be captured by NAPPA.

“interactions dependent on posttranslational modifications may not always be captured.”

Probably not a substantial claim: may not pass S2 (checkable).

The next essential step is to validate the newly identified protein interactions.

“However, the next essential step is to validate the newly identified protein interactions.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Cilia and colleagues examined whether aphid transmission could indirectly affect the interactions between cereal yellow dwarf virus and its plant host.

“Specifically, Cilia et al. assessed how aphid transmission may indirectly modulate the virus–host interactions between the cereal yellow dwarf virus (CYDV) and plants [135].”

• cites [135] sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

Live cell imaging microscopy, tomography, and optogenetics can be integrated with proteomics to yield biological insights that individual techniques cannot provide.

“Additionally, live cell imaging microscopy, tomography, and optogenetics can be further integrated with proteomics to generate biological insights unattainable by individual techniques.”

Probably not a substantial claim: may not pass S2 (checkable).

Proteomics is well suited to complement biochemical, optical, and molecular virology approaches in further clarifying virus-host dynamics.

“Most importantly, proteomics is well suited to complement orthogonal approaches, including biochemical, optical, and molecular virology methods, in the pursuit of further elucidating virus–host dynamics.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Several valuable quantitative MS methods have not yet been applied to studying protein interactions during infection.

“Despite the successful integration of quantitative proteomics with virology, several valuable quantitative MS methods are yet to be applied to studying protein interactions during infection.”

• AssumptionRests on an assumption: Recent years have seen substantial advances in quantitative mass spectrometry-based proteomics.