Targetome Analysis of Malaria Sporozoite Transcription Factor AP2-Sp Reveals Its Role As a Master Regulator

Malaria transmission to humans begins with sporozoite infection of the liver. The elucidation of gene regulation during the sporozoite stage will promote the investigation of mechanisms of liver infection by this parasite and contribute to the development of strategies for preventing malaria transmission. AP2-Sp is a transcription factor (TF) essential for the formation of sporozoites or sporogony, which takes place in oocysts in the midguts of infected mosquitoes. To understand the role of this TF in the transcriptional regulatory system of this stage, we performed chromatin immunoprecipitation sequencing (ChIP-seq) analyses using whole mosquito midguts containing late oocysts as starting material and explored its genome-wide target genes. We identified 697 target genes, comprising those involved in distinct processes parasites experience during this stage, from sporogony to development into the liver stage and representing the majority of genes highly expressed in the sporozoite stage. These results suggest that AP2-Sp determines basal patterns of gene expression by targeting a broad range of genes directly. The ChIP-seq analyses also showed that AP2-Sp maintains its own expression by a transcriptional autoactivation mechanism (positive-feedback loop) and induces all TFs reported to be transcribed at this stage, including AP2-Sp2, AP2-Sp3, and SLARP. The results showed that AP2-Sp exists at the top of the transcriptional cascade of this stage and triggers the formation of this stage as a master regulator. The sporozoite stage plays a central role in malaria transmission from a mosquito to vertebrate host and is an important target for antimalarial strategies. AP2-Sp is a candidate master transcription factor for the sporozoite stage. However, study of its role in gene regulation has been hampered because of difficulties in performing genome-wide studies of gene regulation in this stage. Here, we conquered this problem and revealed that AP2-Sp has the following prominent features as a master transcription factor. First, it determines the repertory of gene expression during this stage. Second, it maintains its own expression through a transcriptional positive-feedback loop and induces all other transcription factors specifically expressed in this stage. This study represents a major breakthrough in fully understanding gene regulation in this important malarial stage.