Graduation Year

2013

Document Type

Dissertation

Degree

D.P.H.

Degree Granting Department

Global Health

Major Professor

John Adams

Abstract

Signal transduction and stage-specific gene expression are essential components of Plasmodium falciparum development. In this study, the putative phosphatase PF13_0027 is investigated as a critical component of intraerythrocytic development contributing to maturation of the late trophozoite. This putative phosphatase was identified during the course of a large-scale insertional mutagenesis project by insertion of the piggyBac (pB) element, containing a human dihydrofolate reductase (hDHFR) drug selection cassette into the open reading frame (ORF) preventing expression and attenuating parasite development. PF13_0027 codes for a protein with a rhodanese (RHD) and dual specificity phosphatase (DUSP) in a tandem arrangement typically identified with mitogen-activated protein kinase (MAPK) phosphatases (MKP). Despite numerous INDELs, the tertiary structure is conserved when compared to the solved structures of MKP homologs. The expression profile reveals transcripts at all stages of the blood cycle with a highest relative abundance in the late trophozoite. Restoration of the phenotype was achieved through genetic complementation using the complete PF13_0027 open reading frame (ORF) under the control of its endogenous promoter. A homology model of PF13_0027 was developed for structural analysis and evaluated using in silico high-throughput screening (HTS) to identify antimalarial compounds with predicted affinity to the active site and used to challenge parasites in vitro. This study reveals that PF13_0027 is a vital component of asexual development and a potential target for a new class of antimalarial compounds targeting phosphorylation pathways in P. falciparum. Discovery of the functional role of this unknown ORF provides additional insight into the importance of MAPK signaling in P. falciparum.

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