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Title: A Novel Secreted Protein, MYR1, Is Central to Toxoplasma ’s Manipulation of Host Cells

Abstract

The intracellular protozoan Toxoplasma gondii dramatically reprograms the transcriptome of host cells it infects, including substantially up-regulating the host oncogene c-myc. By applying a flow cytometry-based selection to infected mouse cells expressing green fluorescent protein fused to c-Myc (c-Myc–GFP), we isolated mutant tachyzoites defective in this host c-Myc up-regulation. Whole-genome sequencing of three such mutants led to the identification of MYR1 (Myc regulation 1; TGGT1_254470) as essential for c-Myc induction. MYR1 is a secreted protein that requires TgASP5 to be cleaved into two stable portions, both of which are ultimately found within the parasitophorous vacuole and at the parasitophorous vacuole membrane. Deletion of MYR1 revealed that in addition to its requirement for c-Myc up-regulation, the MYR1 protein is needed for the ability of Toxoplasma tachyzoites to modulate several other important host pathways, including those mediated by the dense granule effectors GRA16 and GRA24. This result, combined with its location at the parasitophorous vacuole membrane, suggested that MYR1 might be a component of the machinery that translocates Toxoplasma effectors from the parasitophorous vacuole into the host cytosol. Support for this possibility was obtained by showing that transit of GRA24 to the host nucleus is indeed MYR1-dependent. As predicted by this pleiotropicmore » phenotype, parasites deficient in MYR1 were found to be severely attenuated in a mouse model of infection. We conclude, therefore, that MYR1 is a novel protein that plays a critical role in how Toxoplasma delivers effector proteins to the infected host cell and that this is crucial to virulence.« less

Authors:
 [1];  [2];  [2];  [3];  [2];  [2];  [4];  [4];  [3];  [2]
  1. Stanford Univ. School of Medicine, CA (United States); Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
  2. Stanford Univ. School of Medicine, CA (United States)
  3. Univ. of California, Santa Cruz, CA (United States)
  4. Washington Univ., St. Louis, MO (United States)
Publication Date:
Research Org.:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org.:
USDOE; National Institutes of Health (NIH)
OSTI Identifier:
1259517
Grant/Contract Number:  
R21-AI112962; RO1-AI73756; T32-AI007328; P01-35HG000205; T32-AI007290; F31-AI120649; K08AI102946-01; S10RR025518-01; P30-NS069375
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
mBio (Online)
Additional Journal Information:
Journal Volume: 7; Journal Issue: 1; Journal ID: ISSN 2150-7511
Publisher:
American Society for Microbiology
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES

Citation Formats

Franco, Magdalena, Panas, Michael W., Marino, Nicole D., Lee, Mei-Chong Wendy, Buchholz, Kerry R., Kelly, Felice D., Bednarski, Jeffrey J., Sleckman, Barry P., Pourmand, Nader, and Boothroyd, John C. A Novel Secreted Protein, MYR1, Is Central to Toxoplasma ’s Manipulation of Host Cells. United States: N. p., 2016. Web. doi:10.1128/mbio.02231-15.
Franco, Magdalena, Panas, Michael W., Marino, Nicole D., Lee, Mei-Chong Wendy, Buchholz, Kerry R., Kelly, Felice D., Bednarski, Jeffrey J., Sleckman, Barry P., Pourmand, Nader, & Boothroyd, John C. A Novel Secreted Protein, MYR1, Is Central to Toxoplasma ’s Manipulation of Host Cells. United States. doi:10.1128/mbio.02231-15.
Franco, Magdalena, Panas, Michael W., Marino, Nicole D., Lee, Mei-Chong Wendy, Buchholz, Kerry R., Kelly, Felice D., Bednarski, Jeffrey J., Sleckman, Barry P., Pourmand, Nader, and Boothroyd, John C. Tue . "A Novel Secreted Protein, MYR1, Is Central to Toxoplasma ’s Manipulation of Host Cells". United States. doi:10.1128/mbio.02231-15. https://www.osti.gov/servlets/purl/1259517.
@article{osti_1259517,
title = {A Novel Secreted Protein, MYR1, Is Central to Toxoplasma ’s Manipulation of Host Cells},
author = {Franco, Magdalena and Panas, Michael W. and Marino, Nicole D. and Lee, Mei-Chong Wendy and Buchholz, Kerry R. and Kelly, Felice D. and Bednarski, Jeffrey J. and Sleckman, Barry P. and Pourmand, Nader and Boothroyd, John C.},
abstractNote = {The intracellular protozoan Toxoplasma gondii dramatically reprograms the transcriptome of host cells it infects, including substantially up-regulating the host oncogene c-myc. By applying a flow cytometry-based selection to infected mouse cells expressing green fluorescent protein fused to c-Myc (c-Myc–GFP), we isolated mutant tachyzoites defective in this host c-Myc up-regulation. Whole-genome sequencing of three such mutants led to the identification of MYR1 (Myc regulation 1; TGGT1_254470) as essential for c-Myc induction. MYR1 is a secreted protein that requires TgASP5 to be cleaved into two stable portions, both of which are ultimately found within the parasitophorous vacuole and at the parasitophorous vacuole membrane. Deletion of MYR1 revealed that in addition to its requirement for c-Myc up-regulation, the MYR1 protein is needed for the ability of Toxoplasma tachyzoites to modulate several other important host pathways, including those mediated by the dense granule effectors GRA16 and GRA24. This result, combined with its location at the parasitophorous vacuole membrane, suggested that MYR1 might be a component of the machinery that translocates Toxoplasma effectors from the parasitophorous vacuole into the host cytosol. Support for this possibility was obtained by showing that transit of GRA24 to the host nucleus is indeed MYR1-dependent. As predicted by this pleiotropic phenotype, parasites deficient in MYR1 were found to be severely attenuated in a mouse model of infection. We conclude, therefore, that MYR1 is a novel protein that plays a critical role in how Toxoplasma delivers effector proteins to the infected host cell and that this is crucial to virulence.},
doi = {10.1128/mbio.02231-15},
journal = {mBio (Online)},
issn = {2150-7511},
number = 1,
volume = 7,
place = {United States},
year = {2016},
month = {2}
}

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Cited by: 14 works
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Figures / Tables:

FIG 1 FIG 1: Genetic screen to isolate Toxoplasma mutants that fail to induce c-Myc. (A) Flow cytometry histogram of BMMs expressing c-Myc–GFP infected with Toxoplasma wild-type tachyzoites (RH) or Neospora tachyzoites or mock-infected. The gate was set on highly infected cells. The y axis shows relative cell count for each populationmore » (normalized to mode), and the x axis shows green fluorescence intensity. (B) Selection of mutants that fail to induce c-Myc. BMM c-Myc–GFP reporter cells were infected with a mutagenized population of RH tachyzoites and sorted for low GFP fluorescence. This was repeated until a homogeneous population was obtained that induced low GFP fluorescence. The GFP fluorescence profiles for the mutagenized population after 3, 5, and 7 rounds of selection are shown. (Histograms for first two sorts were similar to the one for sort 3 and were omitted.) The profiles for cells infected with wild-type RH and Neospora tachyzoites are shown for comparison. The x and y axes are as described for panel A.« less

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