Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite
Abstract Mutually exclusive expression of the var multigene family is key to immune evasion and pathogenesis in Plasmodium falciparum, but few factors have been shown to play a direct role. We adapted a CRISPR‐based proteomics approach to identify novel factors associated with var genes in their nat...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Springer Nature
2020-08-01
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| Series: | Molecular Systems Biology |
| Subjects: | |
| Online Access: | https://doi.org/10.15252/msb.20209569 |
| _version_ | 1827014816123846656 |
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| author | Jessica M Bryant Sebastian Baumgarten Florent Dingli Damarys Loew Ameya Sinha Aurélie Claës Peter R Preiser Peter C Dedon Artur Scherf |
| author_facet | Jessica M Bryant Sebastian Baumgarten Florent Dingli Damarys Loew Ameya Sinha Aurélie Claës Peter R Preiser Peter C Dedon Artur Scherf |
| author_sort | Jessica M Bryant |
| collection | DOAJ |
| description | Abstract Mutually exclusive expression of the var multigene family is key to immune evasion and pathogenesis in Plasmodium falciparum, but few factors have been shown to play a direct role. We adapted a CRISPR‐based proteomics approach to identify novel factors associated with var genes in their natural chromatin context. Catalytically inactive Cas9 (“dCas9”) was targeted to var gene regulatory elements, immunoprecipitated, and analyzed with mass spectrometry. Known and novel factors were enriched including structural proteins, DNA helicases, and chromatin remodelers. Functional characterization of PfISWI, an evolutionarily divergent putative chromatin remodeler enriched at the var gene promoter, revealed a role in transcriptional activation. Proteomics of PfISWI identified several proteins enriched at the var gene promoter such as acetyl‐CoA synthetase, a putative MORC protein, and an ApiAP2 transcription factor. These findings validate the CRISPR/dCas9 proteomics method and define a new var gene‐associated chromatin complex. This study establishes a tool for targeted chromatin purification of unaltered genomic loci and identifies novel chromatin‐associated factors potentially involved in transcriptional control and/or chromatin organization of virulence genes in the human malaria parasite. |
| first_indexed | 2024-03-07T17:54:29Z |
| format | Article |
| id | doaj.art-1b263eb127434a03a0f52b64bd894d4b |
| institution | Directory Open Access Journal |
| issn | 1744-4292 |
| language | English |
| last_indexed | 2025-02-18T14:14:53Z |
| publishDate | 2020-08-01 |
| publisher | Springer Nature |
| record_format | Article |
| series | Molecular Systems Biology |
| spelling | doaj.art-1b263eb127434a03a0f52b64bd894d4b2024-10-28T09:17:18ZengSpringer NatureMolecular Systems Biology1744-42922020-08-0116812210.15252/msb.20209569Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasiteJessica M Bryant0Sebastian Baumgarten1Florent Dingli2Damarys Loew3Ameya Sinha4Aurélie Claës5Peter R Preiser6Peter C Dedon7Artur Scherf8Biology of Host‐Parasite Interactions Unit, Institut PasteurBiology of Host‐Parasite Interactions Unit, Institut PasteurInstitut Curie, PSL Research University, Centre de Recherche, Mass Spectrometry and Proteomics FacilityInstitut Curie, PSL Research University, Centre de Recherche, Mass Spectrometry and Proteomics FacilitySchool of Biological Sciences, Nanyang Technological UniversityBiology of Host‐Parasite Interactions Unit, Institut PasteurSchool of Biological Sciences, Nanyang Technological UniversityAntimicrobial Resistance Interdisciplinary Research Group, Singapore‐MIT Alliance for Research and TechnologyBiology of Host‐Parasite Interactions Unit, Institut PasteurAbstract Mutually exclusive expression of the var multigene family is key to immune evasion and pathogenesis in Plasmodium falciparum, but few factors have been shown to play a direct role. We adapted a CRISPR‐based proteomics approach to identify novel factors associated with var genes in their natural chromatin context. Catalytically inactive Cas9 (“dCas9”) was targeted to var gene regulatory elements, immunoprecipitated, and analyzed with mass spectrometry. Known and novel factors were enriched including structural proteins, DNA helicases, and chromatin remodelers. Functional characterization of PfISWI, an evolutionarily divergent putative chromatin remodeler enriched at the var gene promoter, revealed a role in transcriptional activation. Proteomics of PfISWI identified several proteins enriched at the var gene promoter such as acetyl‐CoA synthetase, a putative MORC protein, and an ApiAP2 transcription factor. These findings validate the CRISPR/dCas9 proteomics method and define a new var gene‐associated chromatin complex. This study establishes a tool for targeted chromatin purification of unaltered genomic loci and identifies novel chromatin‐associated factors potentially involved in transcriptional control and/or chromatin organization of virulence genes in the human malaria parasite.https://doi.org/10.15252/msb.20209569chromatinCRISPRepigeneticsPlasmodium falciparumvar genes |
| spellingShingle | Jessica M Bryant Sebastian Baumgarten Florent Dingli Damarys Loew Ameya Sinha Aurélie Claës Peter R Preiser Peter C Dedon Artur Scherf Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite Molecular Systems Biology chromatin CRISPR epigenetics Plasmodium falciparum var genes |
| title | Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite |
| title_full | Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite |
| title_fullStr | Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite |
| title_full_unstemmed | Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite |
| title_short | Exploring the virulence gene interactome with CRISPR/dCas9 in the human malaria parasite |
| title_sort | exploring the virulence gene interactome with crispr dcas9 in the human malaria parasite |
| topic | chromatin CRISPR epigenetics Plasmodium falciparum var genes |
| url | https://doi.org/10.15252/msb.20209569 |
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