Journal
Elife
Publication Date
2021
Volume
10
First Page
e73011
Document Type
Open Access Publication
DOI
10.7554/eLife.73011
Rights and Permissions
eLife 2021;10:e73011 DOI: 10.7554/eLife.73011. © 2021, Fu 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
Fu, Yong; Brown, Kevin M.; Jones, Nathaniel G.; Moreno, Silvia N.J.; and Sibley, L. David, "Toxoplasma bradyzoites exhibit physiological plasticity of calcium and energy stores controlling motility and egress." Elife. 10, e73011 (2021).
https://digitalcommons.wustl.edu/open_access_pubs/11193
Figures and figure supplements.pdf (15262 kB)
elife-73011-transrepform1-v3.docx (245 kB)
Transparent reporting form
elife-73011-fig1-data1-v3.xlsx (10 kB)
Figure 1—source data 1 Percentage of egress or release from one vacuole or cyst (related to Figure 1B).
elife-73011-fig1-data2-v3.xlsx (10 kB)
Figure 1—source data 2 Maximum egress distance of egressed or released parasites (related to Figure 1C).
elife-73011-fig1-data3-v3.xlsx (10 kB)
Figure 1—source data 3 Speed of egressed or released parasites (related to Figure 1D).
elife-73011-fig2-data1-v3.xlsx (93 kB)
Figure 2—source data 1 Western blotting analysis of ME49 BAG1-mCherry MIC2-GLuc reporter parasites (related to Figure 2C).
elife-73011-fig2-data2-v3.xlsx (10 kB)
Figure 2—source data 2 Determining MIC2-Gluc secretion by parasites using Gaussia luciferase assay (related to Figure 2D).
elife-73011-fig2-data3-v3.xlsx (14 kB)
Figure 2—source data 3 Monitoring leakage of FNR-mCherry from the parasitophorous vacuole (PV) or cyst matrix following A23187-induced membrane permeabilization (related to Figure 2G).
elife-73011-fig3-data1-v3.xlsx (12 kB)
Figure 3—source data 1 Time of Ca2+ dual reporter tachyzoites and bradyzoites with different ages to Ca2+ peak level in response to A23187 (related to Figure 3C).
elife-73011-fig3-data2-v3.xlsx (25 kB)
Figure 3—source data 2 GCaMP6f fluorescence intensity changes vs. time of GCaMP6f BAG1-mCherry tachyzoites and bradyzoites (related to Figure 3D).
elife-73011-fig3-data3-v3.xlsx (21 kB)
Figure 3—source data 3 Zaprinast-induced Ca2+ responses in GCaMP6f BAG1-mCherry tachyzoites and bradyzoites (related to Figure 3E).
elife-73011-fig3-data4-v3.xlsx (11 kB)
Figure 3—source data 4 Percentage of motile parasites within parasitophorous vacuole or cyst in response to Ca2+ agonists (related to Figure 3G).
elife-73011-fig4-data1-v3.xlsx (15 kB)
Figure 4—source data 1 GCaMP6f fluorescence intensity changes vs. time of GCaMP6f BAG1-mCherry tachyzoites and bradyzoites cultured in C2C12 muscle cells in response to Ca2+ agonists (related to Figure 4C).
elife-73011-fig4-data2-v3.xlsx (11 kB)
Figure 4—source data 2 Time of tachyzoites and bradyzoites cultured in C2C12 muscle cells to reach GCaMP6f fluorescence peak in response to Ca2+ agonists (related to Figure 4D).
elife-73011-fig4-data3-v3.xlsx (11 kB)
Figure 4—source data 3 Percentage of motile parasites within parasitophorous vacuole or cyst cultured in C2C12 muscle cells (related to Figure 4E).
elife-73011-fig4-data4-v3.xlsx (17 kB)
Figure 4—source data 4 Calcium responses of ex vivo cysts in response to Ca2+ agonists (related to Figure 4G).
elife-73011-fig4-data5-v3.xlsx (9 kB)
Figure 4—source data 5 Motility and egress of bradyzoites within ex vivo cysts isolated from chronically infected mice in response to A23187 (related to Figure 4H).
elife-73011-fig5-data1-v3.xlsx (24 kB)
Figure 5—source data 1 Calcium responses of extracellular single tachyzoite and bradyzoite stimulated by Ca2+ agonists (related to Figure 5B).
elife-73011-fig5-data2-v3.xlsx (39 kB)
Figure 5—source data 2 Calcium responses of extracellular GCaMP6f BAG1-mCherry tachyzoites and bradyzoites treated with ionomycin, thapsigargin, and NH4Cl (related to Figure 5D).
elife-73011-fig5-data3-v3.xlsx (29 kB)
Figure 5—source data 3 Calcium responses of extracellular Fluo-8-loaded BAG1-mCherry tachyzoites and bradyzoites treated with ionomycin, thapsigargin, and NH4Cl (related to Figure 5E).
elife-73011-fig6-data1-v3.xlsx (2858 kB)
Figure 6—source data 1 Western blotting analysis of ME49 GCaMP6f-P2A-mTagBFP2 BAG1-mCherry ratiometric reporter parasites (related to Figure 6C).
elife-73011-fig6-data2-v3.xlsx (14 kB)
Figure 6—source data 2 Comparison of Ca2+ calcium basal levels between tachyzoites and bradyzoites using ratiometric reporter (related to Figure 6D).
elife-73011-fig6-data3-v3.xlsx (28 kB)
Figure 6—source data 3 Calcium responses of intracellular GCaMP6f ratiometric reporter tachyzoites and bradyzoites treated with different Ca2+ agonists (related to Figure 6E).
elife-73011-fig6-data4-v3.xlsx (23 kB)
Figure 6—source data 4 Calcium responses of extracellular GCaMP6f ratiometric reporter tachyzoites and bradyzoites treated with different Ca2+ agonists (related to Figure 6F).
elife-73011-fig6-data5-v3.xlsx (13 kB)
Figure 6—source data 5 Comparison of mRNA expression levels of genes encoding calcium-associated channels and pumps in tachyzoites and bradyzoites by qPCR (related to Figure 6G).
elife-73011-fig7-data1-v3.xlsx (12 kB)
Figure 7—source data 1 Calcium responses of GCaMP6f BAG1-mCherry reporter bradyzoites during liberation from in vitro-induced cyst by trypsin (related to Figure 7B).
elife-73011-fig7-data2-v3.xlsx (11 kB)
Figure 7—source data 2 Calcium fluctuation of extracellular single bradyzoites (related to Figure 7D).
elife-73011-fig7-data3-v3.xlsx (15 kB)
Figure 7—source data 3 Effects of inhibition of calcium signaling pathway on the gliding motility of tachyzoites and bradyzoites (related to Figure 7F).
elife-73011-fig8-data1-v3.xlsx (20 kB)
Figure 8—source data 1 Calcium fluorescence fold change of extracellular BAG1-mCherry GCaMP6f dual reporter bradyzoites treated with or without Ca2+ or glucose for 10 min in response to ionomycin (related to Figure 8A).
elife-73011-fig8-data2-v3.xlsx (20 kB)
Figure 8—source data 2 Calcium fluorescence fold change of extracellular BAG1-mCherry GCaMP6f dual reporter bradyzoites treated with or without Ca2+ or glucose for 1 hr in response to ionomycin (related to Figure 8B).
elife-73011-fig8-data3-v3.xlsx (10 kB)
Figure 8—source data 3 Recovery of microneme secretion by bradyzoites in extracellular (EC) buffer containing Ca2+ and glucose (related to Figure 8C).
elife-73011-fig8-data4-v3.xlsx (13 kB)
Figure 8—source data 4 Percentage of gliding motility of bradyzoites treated with or without glucose or Ca2+ for 10 min or 1 hr (related to Figure 8D).
elife-73011-fig8-data5-v3.xlsx (13 kB)
Figure 8—source data 5 Effects of inhibition of adenosine triphosphate (ATP) production pathways on the gliding motility of bradyzoites (related to Figure 8F).
elife-73011-fig8-data6-v3.xlsx (13 kB)
Figure 8—source data 6 High-performance liquid chromatography (HPLC) analysis of adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) levels in bradyzoites treated with glucose and calcium containing extracellular (EC) buffer for 10 min or 1 hr (related to Figure 8G).
elife-73011-fig8-data7-v3.xlsx (10 kB)
Figure 8—source data 7 Adenosine triphosphate (ATP)/adenosine diphosphate (ADP) ratio of bradyzoites treated with glucose and Ca2+ containing extracellular (EC) buffer for 10 min or 1 hr (related to Figure 8H).
elife-73011-fig8-data8-v3.xlsx (10 kB)
Figure 8—source data 8 Energy charge of bradyzoites treated with glucose and Ca2+ containing extracellular (EC) buffer for 10 min or 1 hr (related to Figure 8I).
elife-73011-supp1-v3.xlsx (13 kB)
Supplementary Material
elife-73011-transrepform1-v3.docx (245 kB)
Transparent reporting form
elife-73011-fig1-data1-v3.xlsx (10 kB)
Figure 1—source data 1 Percentage of egress or release from one vacuole or cyst (related to Figure 1B).
elife-73011-fig1-data2-v3.xlsx (10 kB)
Figure 1—source data 2 Maximum egress distance of egressed or released parasites (related to Figure 1C).
elife-73011-fig1-data3-v3.xlsx (10 kB)
Figure 1—source data 3 Speed of egressed or released parasites (related to Figure 1D).
elife-73011-fig2-data1-v3.xlsx (93 kB)
Figure 2—source data 1 Western blotting analysis of ME49 BAG1-mCherry MIC2-GLuc reporter parasites (related to Figure 2C).
elife-73011-fig2-data2-v3.xlsx (10 kB)
Figure 2—source data 2 Determining MIC2-Gluc secretion by parasites using Gaussia luciferase assay (related to Figure 2D).
elife-73011-fig2-data3-v3.xlsx (14 kB)
Figure 2—source data 3 Monitoring leakage of FNR-mCherry from the parasitophorous vacuole (PV) or cyst matrix following A23187-induced membrane permeabilization (related to Figure 2G).
elife-73011-fig3-data1-v3.xlsx (12 kB)
Figure 3—source data 1 Time of Ca2+ dual reporter tachyzoites and bradyzoites with different ages to Ca2+ peak level in response to A23187 (related to Figure 3C).
elife-73011-fig3-data2-v3.xlsx (25 kB)
Figure 3—source data 2 GCaMP6f fluorescence intensity changes vs. time of GCaMP6f BAG1-mCherry tachyzoites and bradyzoites (related to Figure 3D).
elife-73011-fig3-data3-v3.xlsx (21 kB)
Figure 3—source data 3 Zaprinast-induced Ca2+ responses in GCaMP6f BAG1-mCherry tachyzoites and bradyzoites (related to Figure 3E).
elife-73011-fig3-data4-v3.xlsx (11 kB)
Figure 3—source data 4 Percentage of motile parasites within parasitophorous vacuole or cyst in response to Ca2+ agonists (related to Figure 3G).
elife-73011-fig4-data1-v3.xlsx (15 kB)
Figure 4—source data 1 GCaMP6f fluorescence intensity changes vs. time of GCaMP6f BAG1-mCherry tachyzoites and bradyzoites cultured in C2C12 muscle cells in response to Ca2+ agonists (related to Figure 4C).
elife-73011-fig4-data2-v3.xlsx (11 kB)
Figure 4—source data 2 Time of tachyzoites and bradyzoites cultured in C2C12 muscle cells to reach GCaMP6f fluorescence peak in response to Ca2+ agonists (related to Figure 4D).
elife-73011-fig4-data3-v3.xlsx (11 kB)
Figure 4—source data 3 Percentage of motile parasites within parasitophorous vacuole or cyst cultured in C2C12 muscle cells (related to Figure 4E).
elife-73011-fig4-data4-v3.xlsx (17 kB)
Figure 4—source data 4 Calcium responses of ex vivo cysts in response to Ca2+ agonists (related to Figure 4G).
elife-73011-fig4-data5-v3.xlsx (9 kB)
Figure 4—source data 5 Motility and egress of bradyzoites within ex vivo cysts isolated from chronically infected mice in response to A23187 (related to Figure 4H).
elife-73011-fig5-data1-v3.xlsx (24 kB)
Figure 5—source data 1 Calcium responses of extracellular single tachyzoite and bradyzoite stimulated by Ca2+ agonists (related to Figure 5B).
elife-73011-fig5-data2-v3.xlsx (39 kB)
Figure 5—source data 2 Calcium responses of extracellular GCaMP6f BAG1-mCherry tachyzoites and bradyzoites treated with ionomycin, thapsigargin, and NH4Cl (related to Figure 5D).
elife-73011-fig5-data3-v3.xlsx (29 kB)
Figure 5—source data 3 Calcium responses of extracellular Fluo-8-loaded BAG1-mCherry tachyzoites and bradyzoites treated with ionomycin, thapsigargin, and NH4Cl (related to Figure 5E).
elife-73011-fig6-data1-v3.xlsx (2858 kB)
Figure 6—source data 1 Western blotting analysis of ME49 GCaMP6f-P2A-mTagBFP2 BAG1-mCherry ratiometric reporter parasites (related to Figure 6C).
elife-73011-fig6-data2-v3.xlsx (14 kB)
Figure 6—source data 2 Comparison of Ca2+ calcium basal levels between tachyzoites and bradyzoites using ratiometric reporter (related to Figure 6D).
elife-73011-fig6-data3-v3.xlsx (28 kB)
Figure 6—source data 3 Calcium responses of intracellular GCaMP6f ratiometric reporter tachyzoites and bradyzoites treated with different Ca2+ agonists (related to Figure 6E).
elife-73011-fig6-data4-v3.xlsx (23 kB)
Figure 6—source data 4 Calcium responses of extracellular GCaMP6f ratiometric reporter tachyzoites and bradyzoites treated with different Ca2+ agonists (related to Figure 6F).
elife-73011-fig6-data5-v3.xlsx (13 kB)
Figure 6—source data 5 Comparison of mRNA expression levels of genes encoding calcium-associated channels and pumps in tachyzoites and bradyzoites by qPCR (related to Figure 6G).
elife-73011-fig7-data1-v3.xlsx (12 kB)
Figure 7—source data 1 Calcium responses of GCaMP6f BAG1-mCherry reporter bradyzoites during liberation from in vitro-induced cyst by trypsin (related to Figure 7B).
elife-73011-fig7-data2-v3.xlsx (11 kB)
Figure 7—source data 2 Calcium fluctuation of extracellular single bradyzoites (related to Figure 7D).
elife-73011-fig7-data3-v3.xlsx (15 kB)
Figure 7—source data 3 Effects of inhibition of calcium signaling pathway on the gliding motility of tachyzoites and bradyzoites (related to Figure 7F).
elife-73011-fig8-data1-v3.xlsx (20 kB)
Figure 8—source data 1 Calcium fluorescence fold change of extracellular BAG1-mCherry GCaMP6f dual reporter bradyzoites treated with or without Ca2+ or glucose for 10 min in response to ionomycin (related to Figure 8A).
elife-73011-fig8-data2-v3.xlsx (20 kB)
Figure 8—source data 2 Calcium fluorescence fold change of extracellular BAG1-mCherry GCaMP6f dual reporter bradyzoites treated with or without Ca2+ or glucose for 1 hr in response to ionomycin (related to Figure 8B).
elife-73011-fig8-data3-v3.xlsx (10 kB)
Figure 8—source data 3 Recovery of microneme secretion by bradyzoites in extracellular (EC) buffer containing Ca2+ and glucose (related to Figure 8C).
elife-73011-fig8-data4-v3.xlsx (13 kB)
Figure 8—source data 4 Percentage of gliding motility of bradyzoites treated with or without glucose or Ca2+ for 10 min or 1 hr (related to Figure 8D).
elife-73011-fig8-data5-v3.xlsx (13 kB)
Figure 8—source data 5 Effects of inhibition of adenosine triphosphate (ATP) production pathways on the gliding motility of bradyzoites (related to Figure 8F).
elife-73011-fig8-data6-v3.xlsx (13 kB)
Figure 8—source data 6 High-performance liquid chromatography (HPLC) analysis of adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) levels in bradyzoites treated with glucose and calcium containing extracellular (EC) buffer for 10 min or 1 hr (related to Figure 8G).
elife-73011-fig8-data7-v3.xlsx (10 kB)
Figure 8—source data 7 Adenosine triphosphate (ATP)/adenosine diphosphate (ADP) ratio of bradyzoites treated with glucose and Ca2+ containing extracellular (EC) buffer for 10 min or 1 hr (related to Figure 8H).
elife-73011-fig8-data8-v3.xlsx (10 kB)
Figure 8—source data 8 Energy charge of bradyzoites treated with glucose and Ca2+ containing extracellular (EC) buffer for 10 min or 1 hr (related to Figure 8I).
elife-73011-supp1-v3.xlsx (13 kB)
Supplementary Material
