Red blood cell invasion by the malaria parasite is coordinated through recruitment of a bromodomain protein by the PfAP2-I transcription factor. Plasmodium falciparum 3D7

Obligate intracellular parasites must efficiently invade host cells in order to mature and be transmitted. For the malaria parasite Plasmodium falciparum, invasion of host red blood cells (RBCs) is essential. While disrupting RBC invasion by merozoites of the malaria parasite has long been sought as a means to abrogate parasite development, this goal has not been realized. In large part this is because invasion is completed within seconds and most merozoite invasion proteins are functionally redundant. Here we describe a parasite-specific transcription factor, belonging to the Apicomplexan Apetala 2 (ApiAP2) family, responsible for regulating the expression of merozoite genes essential for RBC invasion (PfAP2-I). PfAP2-I binds to a specific DNA motif within the promoters of a large number of invasion genes and is required for their transcription. Although PfAP2-I contains three AP2 DNA-binding domains, we find that only one is essential to blood stage development. In the nucleus, PfAP2-I recruits several chromatin proteins, including Bromodomain Protein 1 (PfBDP1), forming a complex that is required for transcription of the gene targets. Blocking PfAP2-I function would completely prevent RBC invasion and stall progression of the disease thereby representing an optimal new therapeutic target since ApiAP2 proteins are of plant origin and have no counterparts in the human genome. The ChIP assay was performed as described in (Lopez-Rubio, 2012) with some modifications. Briefly, synchronized schizont stage parasite cultures were saponin-lysed until complete red blood cell lysis and formaldehyde-crosslinked. The isolated chromatin was sheared in SDS lysis buffer to obtain a fragment size of 100-150bp. The input and ChIP samples were reverse cross-linked overnight at 45C in the presence of 5M NaCl and purified. Barcoded libraries for Illumina TruSeq single-end sequencing were constructed using NEBNext DNA library preparation reagents (NEB) by following the standard Illumina library preparation protocol. The libraries were PCR-amplified using Kappa HiFi and purified using Agencourt AMPure XP beads. The final libraries were multiplexed with three barcoded samples and 20% PhiX control DNA per lane on an Illumina HiSeq 2000 system to generate 150 base pair single-end reads.