Exploring the virulence gene interactome withCRISPR/dCas9 in the human malaria parasite
Open Access
- 20 August 2020
- journal article
- research article
- Published by Springer Science and Business Media LLC in Molecular Systems Biology
- Vol. 16 (8), e9569
- https://doi.org/10.15252/msb.20209569
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.Keywords
Funding Information
- Institut Pasteur
This publication has 105 references indexed in Scilit:
- Identification of telomere-associated molecules by engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP)Scientific Reports, 2013
- Mechanisms and Functions of ATP-Dependent Chromatin-Remodeling EnzymesCell, 2013
- Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene ExpressionCell, 2013
- Integrative genomics viewerNature Biotechnology, 2011
- Statistical estimation of cell-cycle progression and lineage commitment in Plasmodium falciparum reveals a homogeneous pattern of transcription in ex vivo cultureProceedings of the National Academy of Sciences of the United States of America, 2009
- Improvements to the Percolator Algorithm for Peptide Identification from Shotgun Proteomics Data SetsJournal of Proteome Research, 2009
- Purification of Proteins Associated with Specific Genomic LociCell, 2009
- Mechanisms underlying mutually exclusive expression of virulence genes by malaria parasitesEMBO Reports, 2007
- myProMS, a web server for management and validation of mass spectrometry‐based proteomic dataProteomics, 2007
- Genome sequence of the human malaria parasite Plasmodium falciparumNature, 2002