Abstract
BackgroundPlasmodium falciparum exhibits resistance to the artemisinin component of the frontline antimalarial treatment Artemisinin-based Combination Therapy in South East Asia. Millions of lives will be at risk if artemisinin resistance (ART-R) spreads to Africa. Single non-synonymous mutations in the propeller region of PF3D7_1343700,“K13” are implicated in resistance. In this work, we use transcriptional profiling to characterize a laboratory-generated k13 insertional mutant previously demonstrated to have increased sensitivity to artemisinins to explore the functional role of k13.ResultsA set of RNA-seq and microarray experiments confirmed that the expression profile of k13 is specifically altered during the early ring and early trophozoite stages of the mutant intraerythrocytic development cycle. The down-regulation of k13 transcripts in this mutant during the early ring stage is associated with a transcriptome advance towards a more trophozoite-like state. To discover the specific downstream effect of k13 dysregulation, we developed a new computational method to search for differential gene expression while accounting for the temporal sequence of transcription. We found that the strongest biological signature of the transcriptome shift is an up-regulation of DNA replication and repair genes during the early ring developmental stage and a down-regulation of DNA replication and repair genes during the early trophozoite stage; by contrast, the expressions of housekeeping genes are unchanged. This effect, due to k13 dysregulation, is antagonistic, such that k13 levels are negatively correlated with DNA replication and repair gene expression.ConclusionOur results support a role for k13 as a stress response regulator consistent with the hypothesis that artemisinins mode of action is oxidative stress and k13 as a functional homolog of Keap1 which in humans regulates DNA replication and repair genes in response to oxidative stress.
Highlights
Plasmodium falciparum exhibits resistance to the artemisinin component of the frontline antimalarial treatment Artemisinin-based Combination Therapy in South East Asia
To identify the genes most prominently linked to the disrupted pattern of normal transcription, we developed, what is to our knowledge, a novel computational method to parse out the important differences between the datasets with a temporal sequence called the Dephaser Identifier (DI) algorithm
To identify the most likely regulated transcription factor, we looked for over-representation of dysregulated genes among genes with promoter sequences associated with transcription factors that regulate DNA replication genes described in Campbell et al [30], using Fisher’s exact test (Additional file 5)
Summary
Prior to Artemisinin-based combination therapy (ACT) becoming the World Health Organization recommend treatment for uncomplicated Plasmodium falciparum infection, approximately 1,000,000 people were being killed by malaria annually [2]. Evidence from Mbengue et al [17] suggests that K13 plays a role in regulating ubiquitination This human homolog of K13 is a well-characterized transcriptional regulator of oxidative stress response [18], but the processes regulated by K13 remain unknown in malarial parasites. Birnbaum et al [19] reported that conditionally knocking out k13 halts growth after 3 days at the ring stage, but the mechanisms underlying k13 essentiality are unknown
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