Open AccessExploring the virulence gene interactome with CRISPR / dC as9 in the human malaria parasite
Author(s) -
Bryant Jessica M,
Baumgarten Sebastian,
Dingli Florent,
Loew Damarys,
Sinha Ameya,
Claës Aurélie,
Preiser Peter R,
Dedon Peter C,
Scherf Artur
Publication year - 2020
Publication title -
molecular systems biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 8.523
H-Index - 148
ISSN - 1744-4292
DOI - 10.15252/msb.20209569
Subject(s) - interactome , library science , biology , gene , genetics , computer science
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 (“ dC as9”) 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 Pf ISWI , an evolutionarily divergent putative chromatin remodeler enriched at the var gene promoter, revealed a role in transcriptional activation. Proteomics of Pf ISWI identified several proteins enriched at the var gene promoter such as acetyl‐CoA synthetase, a putative MORC protein, and an Api AP 2 transcription factor. These findings validate the CRISPR / dC as9 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.