Journal
Elife
Publication Date
12-6-2021
Volume
10
First Page
e73137
Document Type
Open Access Publication
DOI
10.7554/eLife.73137
Rights and Permissions
eLife 2021;10:e73137 DOI: 10.7554/eLife.73137. © 2021, Rinkenberger et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
Recommended Citation
Rinkenberger, Nicholas; Abrams, Michael E.; Matta, Sumit K.; Schoggins, John W.; Alto, Neal M.; and Sibley, L, David, "Overexpression screen of interferon-stimulated genes identifies RARRES3 as a restrictor of Toxoplasma gondii infection." Elife. 10, e73137 (2021).
https://digitalcommons.wustl.edu/open_access_pubs/11212
Figures and figure supplements.pdf (7103 kB)
elife-73137-transrepform1-v2.docx (245 kB)
Transparent reporting form
elife-73137-fig1-data1-v2.zip (16 kB)
Figure 1—source data 1 Vacuole size quantitation following CTG-GFP infection of IFN pretreated A549 cells.
elife-73137-fig1-data2-v2.zip (15 kB)
Figure 1—source data 2 Vacuole number quantitation following CTG-GFP infection of IFN pretreated A549 cells.
elife-73137-fig1-data3-v2.zip (15 kB)
Figure 1—source data 3 Quantitation of the percentage of vacuoles containing eight or more parasites following CTG-GFP infection of IFN pretreated A549 cells.
elife-73137-fig1-data4-v2.zip (16 kB)
Figure 1—source data 4 Numerical value summary for the results of the ISG screen.
elife-73137-fig1-data5-v2.zip (10 kB)
Figure 1—source data 5 Quantitation of the total image area infected after CTG-GFP infection of WT or IDO1−/− A549 cells.
elife-73137-fig1-data6-v2.zip (31 kB)
Figure 1—source data 6 Contrast enhanced and labeled actin western blot.
elife-73137-fig1-data7-v2.zip (195 kB)
Figure 1—source data 7 Unmodified actin western blot.
elife-73137-fig1-data8-v2.zip (4 kB)
Figure 1—source data 8 Contrast enhanced and labeled IDO1 western blot.
elife-73137-fig1-data9-v2.zip (142 kB)
Figure 1—source data 9 Unmodified IDO1 western blot.
elife-73137-fig2-data1-v2.zip (15 kB)
Figure 2—source data 1 Cell counts per field for cells ectopically expressing ISG screen hits.
elife-73137-fig2-data2-v2.zip (14 kB)
Figure 2—source data 2 Percentage of A549s ectopically expressing ISG screen hits that stained positive for SYTOX Green.
elife-73137-fig2-data3-v2.zip (21 kB)
Figure 2—source data 3 Number of vacuoles counted after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 36 hr.
elife-73137-fig2-data4-v2.zip (15 kB)
Figure 2—source data 4 Quantitation of vacuole size after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 36 hr. https://cdn.elifesciences.org/articles/73137/elife-73137-fig2-data4-v2.zip
elife-73137-fig2-data5-v2.zip (22 kB)
Figure 2—source data 5 Quantitation of total area infected per sample after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 96 hr.
elife-73137-fig2-data6-v2.zip (13 kB)
Figure 2—source data 6 Quantitation of average infection foci size formed after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 96 hr.
elife-73137-fig3-data1-v2.xlsx (3242 kB)
Figure 3—source data 1 Differential expression of genes after IFNγ treatment relative to mock control.
elife-73137-fig3-data2-v2.xlsx (3252 kB)
Figure 3—source data 2 Differential expression of genes in IRF1 overexpressing cells relative to FLUC expressing control cells.
elife-73137-fig3-data3-v2.xlsx (106 kB)
Figure 3—source data 3 Gene ontology term enrichment for genes significantly induced by IFNγ treatment.
elife-73137-fig3-data4-v2.xlsx (45 kB)
Figure 3—source data 4 Gene ontology term enrichment for genes significantly induced by IRF1 overexpression.
elife-73137-fig4-data1-v2.zip (14 kB)
Figure 4—source data 1 Quantitation of the percentage of IRF1 staining cells after IFNγ treatment of WT or STAT1−/− A549 cells.
elife-73137-fig4-data2-v2.zip (8 kB)
Figure 4—source data 2 Quantitation of the number of vacuoles formed following CTG-GFP infection of RARRES3 or FLUC ectopically expressing STAT1−/− A549 cells.
elife-73137-fig4-data3-v2.zip (7 kB)
Figure 4—source data 3 Vacuole size quantitation following CTG-GFP infection of RARRES3 or FLUC ectopically expressing STAT1−/− A549 cells.
elife-73137-fig4-data4-v2.zip (16 kB)
Figure 4—source data 4 Quantitation of total infected area per sample after infection of STAT1−/− A549s ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig4-data5-v2.zip (6 kB)
Figure 4—source data 5 Quantitation of average infection foci size formed after infection of STAT1−/− A549s ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig4-data6-v2.zip (18 kB)
Figure 4—source data 6 Luminescence values from luciferase assays conducted on lysates from cells expressing a GAS-LUC reporter and RARRES3.
elife-73137-fig4-data7-v2.zip (20 kB)
Figure 4—source data 7 Luminescence values from luciferase assays conducted on lysates from cells expressing a κB-LUC reporter and RARRES3.
elife-73137-fig4-data8-v2.zip (18 kB)
Figure 4—source data 8 Luminescence values from luciferase assays conducted on cell supernatant from cells expressing an ISRE-GLUC reporter and RARRES3.
elife-73137-fig4-data9-v2.zip (17 kB)
Figure 4—source data 9 Luminescence values from luciferase assays conducted on lysates from cells expressing a GFP-LUC reporter control and RARRES3.
elife-73137-fig5-data1-v2.zip (14 kB)
Figure 5—source data 1 Quantitation of total infected area per sample after infection of A549s ectopically expressing WT RARRES3 and mutants with CTG-GFP for 96 hr.
elife-73137-fig5-data2-v2.zip (20 kB)
Figure 5—source data 2 Contrast enhanced and labeled actin western blot.
elife-73137-fig5-data3-v2.zip (256 kB)
Figure 5—source data 3 Unmodified actin western blot.
elife-73137-fig5-data4-v2.zip (25 kB)
Figure 5—source data 4 Contrast enhanced and labeled V5 tag western blot.
elife-73137-fig5-data5-v2.zip (241 kB)
Figure 5—source data 5 Unmodified V5 tag western blot.
elife-73137-fig5-data6-v2.zip (29 kB)
Figure 5—source data 6 Quantitation of total infected area per sample after infection of IFNγ pretreated WT or RARRES3−/− A549s with CTG-GFP for 96 hr.
elife-73137-fig5-data7-v2.zip (15 kB)
Figure 5—source data 7 Quantitation of total infected area per sample after infection of WT or RARRES3−/− A549s ectopically expressing IRF1 with CTG-GFP for 96 hr.
elife-73137-fig6-data1-v2.zip (17 kB)
Figure 6—source data 1 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP for 36 hr.
elife-73137-fig6-data2-v2.zip (22 kB)
Figure 6—source data 2 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP for 36 hr in the presence of Compound 1.
elife-73137-fig6-data3-v2.zip (13 kB)
Figure 6—source data 3 Quantitation of the time of egress in hours for parasites egressing from RARRES3 or FLUC ectopically expressing A549s.
elife-73137-fig6-data4-v2.zip (22 kB)
Figure 6—source data 4 Cell count per field after infection of RARRES3 or FLUC ectopically expressing A549s with CTG-GFP in the presence of Compound 1.
elife-73137-fig6-data5-v2.zip (18 kB)
Figure 6—source data 5 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP for 72 hr in the presence of Compound 1.
elife-73137-fig6-data6-v2.zip (98 kB)
Figure 6—source data 6 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP in the presence of cell death inhibitors.
elife-73137-fig7-data1-v2.zip (7 kB)
Figure 7—source data 1 Quantitation of total infected area per sample after infection of HFFs ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig7-data2-v2.zip (7 kB)
Figure 7—source data 2 Quantitation of average number of infection foci formed after infection of HFFs ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig7-data3-v2.zip (26 kB)
Figure 7—source data 3 Absorbance values for lactate dehydrogenase activity assays using supernatants from IFNγ pretreated WT or RARRES3−/− HFF cells infected with CTG-GFP.
elife-73137-fig7-data4-v2.zip (17 kB)
Figure 7—source data 4 Quantitation of total infected area per sample after infection of IFNγ pretreated WT or RARRES3−/− HFFs ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig7-data5-v2.zip (26 kB)
Figure 7—source data 5 Absorbance values for lactate dehydrogenase activity assays using supernatants from IFNγ pretreated HFF cells infected with CTG-GFP in the presence of Compound 1.
elife-73137-video1.mp4 (5902 kB)
Live imaging of FLUC control ectopically expressing A549s infected with CTG-GFP in 15 min intervals. A549 cells were transduced with TRIP.FLUC control and infected 72 hr later with CTG-GFP. Live infection was imaged every 15 min starting 12 hr postinfection until 50 hr postinfection.
elife-73137-video2.mp4 (5325 kB)
Live imaging of RARRES3 ectopically expressing A549s infected with CTG-GFP in 15 min intervals. A549 cells were transduced with TRIP.RARRES3 and infected 72 hr later with CTG-GFP. Live infection was imaged every 15 min starting 12 hr postinfection until 50 hr postinfection.
elife-73137-supp1-v2.xlsx (123 kB)
Supplementary file 1 Summary of overexpression screen in A549 cells.
elife-73137-supp2-v2.xlsx (10823 kB)
Supplementary file 2 List of genes induced by IRF1 ectopic expression or IFNγ treatment in A549 cells.
elife-73137-supp3-v2.xlsx (23 kB)
Supplementary file 3 Comparison of genes induced by IRF1 or IFNγ with genes in the type II ISG screen library.
elife-73137-supp4-v2.docx (13 kB)
Supplementary file 4 List of primer sets used.
elife-73137-supp5-v2.zip (957 kB)
Source data 1 GFP channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing FLUC control.
elife-73137-supp6-v2.zip (4789 kB)
Source data 2 Phase channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing FLUC control.
elife-73137-supp7-v2.zip (1145 kB)
Source data 3 GFP channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing RARRES3 control.
elife-73137-supp8-v2.zip (4813 kB)
Source data 4 Phase channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing RARRES3 control.
elife-73137-transrepform1-v2.docx (245 kB)
Transparent reporting form
elife-73137-fig1-data1-v2.zip (16 kB)
Figure 1—source data 1 Vacuole size quantitation following CTG-GFP infection of IFN pretreated A549 cells.
elife-73137-fig1-data2-v2.zip (15 kB)
Figure 1—source data 2 Vacuole number quantitation following CTG-GFP infection of IFN pretreated A549 cells.
elife-73137-fig1-data3-v2.zip (15 kB)
Figure 1—source data 3 Quantitation of the percentage of vacuoles containing eight or more parasites following CTG-GFP infection of IFN pretreated A549 cells.
elife-73137-fig1-data4-v2.zip (16 kB)
Figure 1—source data 4 Numerical value summary for the results of the ISG screen.
elife-73137-fig1-data5-v2.zip (10 kB)
Figure 1—source data 5 Quantitation of the total image area infected after CTG-GFP infection of WT or IDO1−/− A549 cells.
elife-73137-fig1-data6-v2.zip (31 kB)
Figure 1—source data 6 Contrast enhanced and labeled actin western blot.
elife-73137-fig1-data7-v2.zip (195 kB)
Figure 1—source data 7 Unmodified actin western blot.
elife-73137-fig1-data8-v2.zip (4 kB)
Figure 1—source data 8 Contrast enhanced and labeled IDO1 western blot.
elife-73137-fig1-data9-v2.zip (142 kB)
Figure 1—source data 9 Unmodified IDO1 western blot.
elife-73137-fig2-data1-v2.zip (15 kB)
Figure 2—source data 1 Cell counts per field for cells ectopically expressing ISG screen hits.
elife-73137-fig2-data2-v2.zip (14 kB)
Figure 2—source data 2 Percentage of A549s ectopically expressing ISG screen hits that stained positive for SYTOX Green.
elife-73137-fig2-data3-v2.zip (21 kB)
Figure 2—source data 3 Number of vacuoles counted after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 36 hr.
elife-73137-fig2-data4-v2.zip (15 kB)
Figure 2—source data 4 Quantitation of vacuole size after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 36 hr. https://cdn.elifesciences.org/articles/73137/elife-73137-fig2-data4-v2.zip
elife-73137-fig2-data5-v2.zip (22 kB)
Figure 2—source data 5 Quantitation of total area infected per sample after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 96 hr.
elife-73137-fig2-data6-v2.zip (13 kB)
Figure 2—source data 6 Quantitation of average infection foci size formed after infection of A549s ectopically expressing ISG screen hits with CTG-GFP for 96 hr.
elife-73137-fig3-data1-v2.xlsx (3242 kB)
Figure 3—source data 1 Differential expression of genes after IFNγ treatment relative to mock control.
elife-73137-fig3-data2-v2.xlsx (3252 kB)
Figure 3—source data 2 Differential expression of genes in IRF1 overexpressing cells relative to FLUC expressing control cells.
elife-73137-fig3-data3-v2.xlsx (106 kB)
Figure 3—source data 3 Gene ontology term enrichment for genes significantly induced by IFNγ treatment.
elife-73137-fig3-data4-v2.xlsx (45 kB)
Figure 3—source data 4 Gene ontology term enrichment for genes significantly induced by IRF1 overexpression.
elife-73137-fig4-data1-v2.zip (14 kB)
Figure 4—source data 1 Quantitation of the percentage of IRF1 staining cells after IFNγ treatment of WT or STAT1−/− A549 cells.
elife-73137-fig4-data2-v2.zip (8 kB)
Figure 4—source data 2 Quantitation of the number of vacuoles formed following CTG-GFP infection of RARRES3 or FLUC ectopically expressing STAT1−/− A549 cells.
elife-73137-fig4-data3-v2.zip (7 kB)
Figure 4—source data 3 Vacuole size quantitation following CTG-GFP infection of RARRES3 or FLUC ectopically expressing STAT1−/− A549 cells.
elife-73137-fig4-data4-v2.zip (16 kB)
Figure 4—source data 4 Quantitation of total infected area per sample after infection of STAT1−/− A549s ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig4-data5-v2.zip (6 kB)
Figure 4—source data 5 Quantitation of average infection foci size formed after infection of STAT1−/− A549s ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig4-data6-v2.zip (18 kB)
Figure 4—source data 6 Luminescence values from luciferase assays conducted on lysates from cells expressing a GAS-LUC reporter and RARRES3.
elife-73137-fig4-data7-v2.zip (20 kB)
Figure 4—source data 7 Luminescence values from luciferase assays conducted on lysates from cells expressing a κB-LUC reporter and RARRES3.
elife-73137-fig4-data8-v2.zip (18 kB)
Figure 4—source data 8 Luminescence values from luciferase assays conducted on cell supernatant from cells expressing an ISRE-GLUC reporter and RARRES3.
elife-73137-fig4-data9-v2.zip (17 kB)
Figure 4—source data 9 Luminescence values from luciferase assays conducted on lysates from cells expressing a GFP-LUC reporter control and RARRES3.
elife-73137-fig5-data1-v2.zip (14 kB)
Figure 5—source data 1 Quantitation of total infected area per sample after infection of A549s ectopically expressing WT RARRES3 and mutants with CTG-GFP for 96 hr.
elife-73137-fig5-data2-v2.zip (20 kB)
Figure 5—source data 2 Contrast enhanced and labeled actin western blot.
elife-73137-fig5-data3-v2.zip (256 kB)
Figure 5—source data 3 Unmodified actin western blot.
elife-73137-fig5-data4-v2.zip (25 kB)
Figure 5—source data 4 Contrast enhanced and labeled V5 tag western blot.
elife-73137-fig5-data5-v2.zip (241 kB)
Figure 5—source data 5 Unmodified V5 tag western blot.
elife-73137-fig5-data6-v2.zip (29 kB)
Figure 5—source data 6 Quantitation of total infected area per sample after infection of IFNγ pretreated WT or RARRES3−/− A549s with CTG-GFP for 96 hr.
elife-73137-fig5-data7-v2.zip (15 kB)
Figure 5—source data 7 Quantitation of total infected area per sample after infection of WT or RARRES3−/− A549s ectopically expressing IRF1 with CTG-GFP for 96 hr.
elife-73137-fig6-data1-v2.zip (17 kB)
Figure 6—source data 1 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP for 36 hr.
elife-73137-fig6-data2-v2.zip (22 kB)
Figure 6—source data 2 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP for 36 hr in the presence of Compound 1.
elife-73137-fig6-data3-v2.zip (13 kB)
Figure 6—source data 3 Quantitation of the time of egress in hours for parasites egressing from RARRES3 or FLUC ectopically expressing A549s.
elife-73137-fig6-data4-v2.zip (22 kB)
Figure 6—source data 4 Cell count per field after infection of RARRES3 or FLUC ectopically expressing A549s with CTG-GFP in the presence of Compound 1.
elife-73137-fig6-data5-v2.zip (18 kB)
Figure 6—source data 5 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP for 72 hr in the presence of Compound 1.
elife-73137-fig6-data6-v2.zip (98 kB)
Figure 6—source data 6 Absorbance values for lactate dehydrogenase activity assays using supernatants from RARRES3 or FLUC ectopically expressing A549 cells infected with CTG-GFP in the presence of cell death inhibitors.
elife-73137-fig7-data1-v2.zip (7 kB)
Figure 7—source data 1 Quantitation of total infected area per sample after infection of HFFs ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig7-data2-v2.zip (7 kB)
Figure 7—source data 2 Quantitation of average number of infection foci formed after infection of HFFs ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig7-data3-v2.zip (26 kB)
Figure 7—source data 3 Absorbance values for lactate dehydrogenase activity assays using supernatants from IFNγ pretreated WT or RARRES3−/− HFF cells infected with CTG-GFP.
elife-73137-fig7-data4-v2.zip (17 kB)
Figure 7—source data 4 Quantitation of total infected area per sample after infection of IFNγ pretreated WT or RARRES3−/− HFFs ectopically expressing RARRES3 or FLUC with CTG-GFP for 96 hr.
elife-73137-fig7-data5-v2.zip (26 kB)
Figure 7—source data 5 Absorbance values for lactate dehydrogenase activity assays using supernatants from IFNγ pretreated HFF cells infected with CTG-GFP in the presence of Compound 1.
elife-73137-video1.mp4 (5902 kB)
Live imaging of FLUC control ectopically expressing A549s infected with CTG-GFP in 15 min intervals. A549 cells were transduced with TRIP.FLUC control and infected 72 hr later with CTG-GFP. Live infection was imaged every 15 min starting 12 hr postinfection until 50 hr postinfection.
elife-73137-video2.mp4 (5325 kB)
Live imaging of RARRES3 ectopically expressing A549s infected with CTG-GFP in 15 min intervals. A549 cells were transduced with TRIP.RARRES3 and infected 72 hr later with CTG-GFP. Live infection was imaged every 15 min starting 12 hr postinfection until 50 hr postinfection.
elife-73137-supp1-v2.xlsx (123 kB)
Supplementary file 1 Summary of overexpression screen in A549 cells.
elife-73137-supp2-v2.xlsx (10823 kB)
Supplementary file 2 List of genes induced by IRF1 ectopic expression or IFNγ treatment in A549 cells.
elife-73137-supp3-v2.xlsx (23 kB)
Supplementary file 3 Comparison of genes induced by IRF1 or IFNγ with genes in the type II ISG screen library.
elife-73137-supp4-v2.docx (13 kB)
Supplementary file 4 List of primer sets used.
elife-73137-supp5-v2.zip (957 kB)
Source data 1 GFP channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing FLUC control.
elife-73137-supp6-v2.zip (4789 kB)
Source data 2 Phase channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing FLUC control.
elife-73137-supp7-v2.zip (1145 kB)
Source data 3 GFP channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing RARRES3 control.
elife-73137-supp8-v2.zip (4813 kB)
Source data 4 Phase channel of live video microscopy for CTG-GFP infection in A549 cells ectopically expressing RARRES3 control.
