Abstract
SummaryA major obstacle in analyzing gene function in apicomplexan parasites is the absence of a practical regulatable expression system. Here, we identified functional transcriptional activation domains within Apicomplexan AP2 (ApiAP2) family transcription factors. These ApiAP2 transactivation domains were validated in blood-, liver-, and mosquito-stage parasites and used to create a robust conditional expression system for stage-specific, tetracycline-dependent gene regulation in Toxoplasma gondii, Plasmodium berghei, and Plasmodium falciparum. To demonstrate the utility of this system, we created conditional knockdowns of two essential P. berghei genes: profilin (PRF), a protein implicated in parasite invasion, and N-myristoyltransferase (NMT), which catalyzes protein acylation. Tetracycline-induced repression of PRF and NMT expression resulted in a dramatic reduction in parasite viability. This efficient regulatable system will allow for the functional characterization of essential proteins that are found in these important parasites.
Highlights
Access to a generalizable system that tightly controls gene expression has been long sought after in molecular malaria research
The adaptation of TATi-2 to Plasmodium, successful in tightly controlling transgene expression from multicopy episomal plasmids (Meissner et al, 2005), failed to find utility for the construction of conditional knockouts both in P. falciparum and P. berghei, probably due to insufficient transactivation activity
All proof-of-principle data for the prf and nmt genes were obtained using TRAD4 in P. berghei, this study provides a panel of transactivation sequences of varying strengths, which will be useful to achieve tailored levels of regulatable gene expression
Summary
A major obstacle in analyzing gene function in apicomplexan parasites is the absence of a practical regulatable expression system. We identified functional transcriptional activation domains within Apicomplexan AP2 (ApiAP2) family transcription factors. These ApiAP2 transactivation domains were validated in blood-, liver-, and mosquito-stage parasites and used to create a robust conditional expression system for stage-specific, tetracyclinedependent gene regulation in Toxoplasma gondii, Plasmodium berghei, and Plasmodium falciparum. Tetracycline-induced repression of PRF and NMT expression resulted in a dramatic reduction in parasite viability. This efficient regulatable system will allow for the functional characterization of essential proteins that are found in these important parasites
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
