2A and the auxin-based degron system facilitate control of protein levels in Plasmodium falciparum

Analysis of gene function in Plasmodium falciparum, the most important human malaria parasite, is restricted by the lack of robust and simple reverse genetic tools. Approaches to manipulate protein levels post-translationally are powerful tools to study protein-off effects especially in the haploid...

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Bibliographic Details
Published in:PloS one 2013-11, Vol.8 (11), p.e78661
Main Authors: Kreidenweiss, Andrea, Hopkins, Annika V, Mordmüller, Benjamin
Format: Article
Language:English
Subjects:
Analysis
Bacterial Proteins - metabolism
Blood & organ donations
Cloning
Control systems
Cysteine Endopeptidases - biosynthesis
Cysteine Endopeptidases - genetics
Erythrocytes
Fluorescence
Foot & mouth disease
Foot-and-mouth disease
Gene expression
Genes
Genetic aspects
Genetic engineering
Genetic vectors
Genomes
HeLa Cells
Humans
Indoleacetic Acids - pharmacology
Inhibitory Concentration 50
Laboratories
Luminescent Proteins - metabolism
Malaria
Parasites
Plasmodium falciparum
Plasmodium falciparum - drug effects
Plasmodium falciparum - metabolism
Post-translation
Proteins
Proteolysis - drug effects
Protozoan Proteins - metabolism
Transcriptional Activation
Vector-borne diseases
Viability
Viral Proteins - biosynthesis
Viral Proteins - genetics
Viruses
Yellow fluorescent protein
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Summary:Analysis of gene function in Plasmodium falciparum, the most important human malaria parasite, is restricted by the lack of robust and simple reverse genetic tools. Approaches to manipulate protein levels post-translationally are powerful tools to study protein-off effects especially in the haploid malaria parasite where genetic knockouts of essential genes are lethal. We investigated if the auxin-inducible degron system is functional in P. falciparum and found that degron-tagged yellow fluorescent protein levels were efficiently reduced upon addition of auxin which otherwise had no effect on parasite viability. The genetic components required in this conditional approach were co-expressed in P. falciparum by applying the small peptide 2A. 2A is a self-processing peptide from Foot-And-Mouth Disease virus that allows the whole conditional system to be accommodated on a single plasmid vector and ensures stoichiometric expression levels.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0078661