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Quantitative temporal viromics: an approach to investigate host-pathogen interaction.

Weekes MP, Tomasec P, Huttlin EL, Fielding CA, Nusinow D, Stanton RJ, Wang EC, Aicheler R, Murrell I, Wilkinson GW, Lehner PJ, Gygi SP - Cell (2014)

Bottom Line: QTV predicted natural killer and T cell ligands, as well as 29 viral proteins present at the cell surface, potential therapeutic targets.Temporal profiles of >80% of HCMV canonical genes and 14 noncanonical HCMV open reading frames were defined.QTV is a powerful method that can yield important insights into viral infection and is applicable to any virus with a robust in vitro model.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA; Cambridge Institute for Medical Research, University of Cambridge, Hills Road, Cambridge CB2 0XY, UK. Electronic address: mpw1001@cam.ac.uk.

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Definition of Temporal Classes of HCMV Gene Expression(A) The k-means method was used to cluster all quantified HCMV proteins (experiment WCL2) into four or five classes. Shown are the average temporal profiles of each class. With four classes, proteins grouped into classes similar to the classical IE/E/E-L/L cascades. With five classes, a distinct temporal profile appeared (blue).(B) Number of temporal classes of HCMV gene expression. The summed distance of each protein from its cluster centroid was calculated for one to 14 classes and plotted. The point of inflexion fell between five and seven classes.(C) Top: temporal profiles of proteins in each k-means class (experiment WCL2) were subjected to hierarchical clustering by Euclidian distance. Both UL112 and isoform p50 of UL112 (UL112-2) were quantified. UL122 was excluded from clustering due to uncertainty in peptide assignment. Bottom: experiment WCL3. Viral protein profiles were further assessed in the presence or absence of the viral DNA replication inhibitor PFA. Proteins are displayed in the same order as the clusters defined in the upper panels.(D) Temporal profiles of typical proteins from each cluster (upper panel), and the corresponding profiles in the presence or absence of PFA (lower panel).See also Figure S5, Table S6, and Data S1.
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fig5: Definition of Temporal Classes of HCMV Gene Expression(A) The k-means method was used to cluster all quantified HCMV proteins (experiment WCL2) into four or five classes. Shown are the average temporal profiles of each class. With four classes, proteins grouped into classes similar to the classical IE/E/E-L/L cascades. With five classes, a distinct temporal profile appeared (blue).(B) Number of temporal classes of HCMV gene expression. The summed distance of each protein from its cluster centroid was calculated for one to 14 classes and plotted. The point of inflexion fell between five and seven classes.(C) Top: temporal profiles of proteins in each k-means class (experiment WCL2) were subjected to hierarchical clustering by Euclidian distance. Both UL112 and isoform p50 of UL112 (UL112-2) were quantified. UL122 was excluded from clustering due to uncertainty in peptide assignment. Bottom: experiment WCL3. Viral protein profiles were further assessed in the presence or absence of the viral DNA replication inhibitor PFA. Proteins are displayed in the same order as the clusters defined in the upper panels.(D) Temporal profiles of typical proteins from each cluster (upper panel), and the corresponding profiles in the presence or absence of PFA (lower panel).See also Figure S5, Table S6, and Data S1.

Mentions: Most prior studies of the temporal expression of canonical HCMV proteins have employed immunoblots at only one or a few time points. Comparison between different reports has been complicated by lack of specific antibodies and study-to-study variation in the HCMV strain used, with laboratory-adapted strains AD169 and Towne (which lack a large number of ORFs as a result of extensive passage in vitro) often used in preference to isolates containing a complete genome (Ma et al., 2012). We were able to quantify the temporal expression of the majority (139/171) of the canonical HCMV proteins as well as 14 noncanonical ORFs; most of these proteins were quantified in a single experiment (Figure 5, plots for all quantified viral proteins in Data S1).


Quantitative temporal viromics: an approach to investigate host-pathogen interaction.

Weekes MP, Tomasec P, Huttlin EL, Fielding CA, Nusinow D, Stanton RJ, Wang EC, Aicheler R, Murrell I, Wilkinson GW, Lehner PJ, Gygi SP - Cell (2014)

Definition of Temporal Classes of HCMV Gene Expression(A) The k-means method was used to cluster all quantified HCMV proteins (experiment WCL2) into four or five classes. Shown are the average temporal profiles of each class. With four classes, proteins grouped into classes similar to the classical IE/E/E-L/L cascades. With five classes, a distinct temporal profile appeared (blue).(B) Number of temporal classes of HCMV gene expression. The summed distance of each protein from its cluster centroid was calculated for one to 14 classes and plotted. The point of inflexion fell between five and seven classes.(C) Top: temporal profiles of proteins in each k-means class (experiment WCL2) were subjected to hierarchical clustering by Euclidian distance. Both UL112 and isoform p50 of UL112 (UL112-2) were quantified. UL122 was excluded from clustering due to uncertainty in peptide assignment. Bottom: experiment WCL3. Viral protein profiles were further assessed in the presence or absence of the viral DNA replication inhibitor PFA. Proteins are displayed in the same order as the clusters defined in the upper panels.(D) Temporal profiles of typical proteins from each cluster (upper panel), and the corresponding profiles in the presence or absence of PFA (lower panel).See also Figure S5, Table S6, and Data S1.
© Copyright Policy - CC BY
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4048463&req=5

fig5: Definition of Temporal Classes of HCMV Gene Expression(A) The k-means method was used to cluster all quantified HCMV proteins (experiment WCL2) into four or five classes. Shown are the average temporal profiles of each class. With four classes, proteins grouped into classes similar to the classical IE/E/E-L/L cascades. With five classes, a distinct temporal profile appeared (blue).(B) Number of temporal classes of HCMV gene expression. The summed distance of each protein from its cluster centroid was calculated for one to 14 classes and plotted. The point of inflexion fell between five and seven classes.(C) Top: temporal profiles of proteins in each k-means class (experiment WCL2) were subjected to hierarchical clustering by Euclidian distance. Both UL112 and isoform p50 of UL112 (UL112-2) were quantified. UL122 was excluded from clustering due to uncertainty in peptide assignment. Bottom: experiment WCL3. Viral protein profiles were further assessed in the presence or absence of the viral DNA replication inhibitor PFA. Proteins are displayed in the same order as the clusters defined in the upper panels.(D) Temporal profiles of typical proteins from each cluster (upper panel), and the corresponding profiles in the presence or absence of PFA (lower panel).See also Figure S5, Table S6, and Data S1.
Mentions: Most prior studies of the temporal expression of canonical HCMV proteins have employed immunoblots at only one or a few time points. Comparison between different reports has been complicated by lack of specific antibodies and study-to-study variation in the HCMV strain used, with laboratory-adapted strains AD169 and Towne (which lack a large number of ORFs as a result of extensive passage in vitro) often used in preference to isolates containing a complete genome (Ma et al., 2012). We were able to quantify the temporal expression of the majority (139/171) of the canonical HCMV proteins as well as 14 noncanonical ORFs; most of these proteins were quantified in a single experiment (Figure 5, plots for all quantified viral proteins in Data S1).

Bottom Line: QTV predicted natural killer and T cell ligands, as well as 29 viral proteins present at the cell surface, potential therapeutic targets.Temporal profiles of >80% of HCMV canonical genes and 14 noncanonical HCMV open reading frames were defined.QTV is a powerful method that can yield important insights into viral infection and is applicable to any virus with a robust in vitro model.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA; Cambridge Institute for Medical Research, University of Cambridge, Hills Road, Cambridge CB2 0XY, UK. Electronic address: mpw1001@cam.ac.uk.

Show MeSH
Related in: MedlinePlus