PfAP2Tel, harbouring a non-canonical DNA-binding AP2 domain, binds toPlasmodium falciparumtelomeres

Abstract

The telomeres of the malaria parasite Plasmodium falciparum are essential not only for chromosome end maintenance during blood stage development in humans but also to generate genetic diversity by facilitating homologous recombination of subtelomeric, multigene virulence families such as var and rifin. However, other than the telomerase PfTERT, proteins that act at P. falciparum telomeres are poorly characterised. To isolate components that bind to telomeres, we performed oligonucleotide pulldowns and electromobility shift assays with a telomeric DNA probe and identified a non-canonical member of the ApiAP2 family of transcription factors, PfAP2Tel (encoded by PF3D7_0622900), as a component of the P. falciparum telomere-binding protein complex. PfAP2Tel is expressed throughout the intra-erythrocytic life cycle and localises to the nuclear periphery, co-localising with telomeric clusters. Furthermore, EMSAs using the recombinant protein demonstrated direct binding of PfAP2Tel to telomeric repeats in vitro, while genome-wide chromatin immunoprecipitation followed by next generation sequencing corroborated the high specificity of this protein to telomeric ends of all 14 chromosomes in vivo. Taken together, our data describe a novel function for ApiAP2 proteins at chromosome ends and open new avenues to study the molecular machinery that regulates telomere function in P. falciparum.

Characterization of the accessible genome in the human malaria parasite Plasmodium falciparum

“…The more distal enhancer-like regions are commonly found at the ends of chromosomes, occupying subtelomeric repeats such as the TAREs and sometimes very far from the nearest gene. This is likely because TAREs are highly transcribed (and thus presumably contain promoters and enhancers) ( 42 , 83 ). Interestingly, we find that 22 subtelomeric var genes have enhancer-like THSS regions.…”

“…As a validation for our strategy to investigate TF binding events in vivo , we report the overlap between the ATAC-seq peaks and the binding sites of the TFs: AP2Tel, SIP2, AP2-I and other proteins such as BDP1, which have been determined experimentally ( 35 , 41 , 42 , 62 ) ( Supplementary Table S6 ). In the case of AP2-I, we find that 86% of the binding sites intersect with ATAC-seq peaks ( Supplementary Table S6 ).…”

“…Tracks show normalized/input-corrected ATAC-seq signal. The location of known TSSs ( 48 , 66 ) and the binding sites for SIP2 and AP2Tel ( 35 , 42 ) are included. All tracks are shown at equal scale.…”

“…Accordingly, the genes encoding these TFs are expressed in a stage-specific manner ( 45 , 46 ). The cognate motifs of several ApiAP2 TFs have been determined experimentally ( 47 ), but only a few of these have been tested in vivo by chromatin immunoprecipitation-sequencing (ChIP-seq) ( 41 , 42 ), mainly because of the difficulties in obtaining antibodies against Plasmodium proteins ( 47 ). Profiling of accessible chromatin can be used to reveal gene regulatory networks and to identify novel trans-acting factors and cis -regulatory elements.…”

Real-time in vivo Global Transcriptional Dynamics During Plasmodium falciparum Blood-stage Development

“…To date, only a single family of specific transcription factors, the 27-member Apicomplexan AP2 (ApiAP2) proteins, has emerged as transcriptional regulators with functions across all developmental stages 15,[17][18][19][20][21][22][23][24][25] .…”