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.

Details

Title
Exploring the virulence gene interactome with CRISPR / dC as9 in the human malaria parasite
Author
Bryant, Jessica M 1   VIAFID ORCID Logo  ; Baumgarten, Sebastian 1   VIAFID ORCID Logo  ; Dingli, Florent 2   VIAFID ORCID Logo  ; Loew, Damarys 2   VIAFID ORCID Logo  ; Sinha, Ameya 3   VIAFID ORCID Logo  ; Claës, Aurélie 1 ; Preiser, Peter R 3   VIAFID ORCID Logo  ; Dedon, Peter C 4   VIAFID ORCID Logo  ; Scherf, Artur 1   VIAFID ORCID Logo 

 Biology of Host‐Parasite Interactions Unit, Institut Pasteur, Paris, France; INSERM U1201, Paris, France; CNRS ERL9195, Paris, France 
 Institut Curie, PSL Research University, Centre de Recherche, Mass Spectrometry and Proteomics Facility, Paris, France 
 School of Biological Sciences, Nanyang Technological University, Singapore, Singapore; Antimicrobial Resistance Interdisciplinary Research Group, Singapore‐MIT Alliance for Research and Technology, Singapore, Singapore 
 Antimicrobial Resistance Interdisciplinary Research Group, Singapore‐MIT Alliance for Research and Technology, Singapore, Singapore; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA 
Section
Articles
Publication year
2020
Publication date
Aug 2020
Publisher
EMBO Press
e-ISSN
17444292
Source type
Scholarly Journal
Language of publication
English
ProQuest document ID
2494307355
Copyright
© 2020. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.