Biology subjects

Llinas, M.

Publications and source records attributed to Llinas, M..

At least 19 recordsLinked to original sources

bioRxiv2026

A Multidimensional Analysis of the Bimodal Piperaquine Response in Plasmodium falciparum

Malaria remains a pressing global health challenge, with the continued emergence of resistance threatening the long-term efficacy of artemisinin-based combination therapies (ACTs). Piperaquine (PPQ), an important partner drug in artemisinin-based combination therapies exhibits a unique bimodal dose-response phenotype associated with reduced susceptibility, yet the biological mechanism underlying this phenotype remains unknown. This phenotype is strongly associated with mutations in pfcrt and copy number amplification of plasmepsin II/III (pm II/III). Given that plasmepsins play a central role in hemoglobin degradation within the blood stage parasite digestive vacuole, and that PPQ accumulates within this compartment and perturbs heme detoxification, this phenotype likely reflects alterations in fundamental biological processes alongside drug-specific effects. We used isogenic PPQ-resistant parasite clones differing only in pm II/III copy number to integrate phenotypes with metabolic changes, and transcriptional responses to ascertain the impact of genotype combinations on parasite response to PPQ. Across increasing PPQ concentrations, parasites with elevated pm II/III copy number exhibited distinct metabolic responses compared to single-copy parasites, specifically, an altered abundance of peptides derived from hemoglobin degradation, directly implicating a core biological pathway long associated with plasmepsin function. The combination of metabolic and transcriptional data with phenotypic measurements supports a model in which increased plasmepsin expression enhances the parasites capacity to sustain hemoglobin digestion and associated metabolic activity under high PPQ concentrations. This points to a mechanistic basis for continued parasite survival, indicating that changes in hemoglobin processing within the digestive vacuole contribute to the bimodal response to PPQ. Molecular dynamics simulations further support a direct interaction between PPQ and PM II/III, as a mechanism by which these proteins impact PPQ response dynamics through both modulation of hemoglobin digestion and protein-drug interactions within the digestive vacuole.

molecular biology↗

bioRxiv2026

The CCCH-type zinc-finger PfMD3 promotes translation for malaria parasite transmission

Sexual maturation and fertility of the malaria causing parasite, Plasmodium spp., is critical to transmission between hosts. This developmental process involves an intricate gene expression regulatory network that drives gametocyte commitment, development, and the production of male and female gametes. Here, we characterized the role of the CCCH-type zinc-finger protein Plasmodium falciparum Male Development 3 (PfMD3) in the formation of gametes. Genetic disruption of PfMD3 revealed a male fertility defect that decreased infectivity to mosquitoes. Molecular characterization of PfMD3 revealed its binding to mRNA transcripts associated with male gamete development through a specific RNA sequence. Gametocyte proteins encoded by these PfMD3-bound transcripts decreased significantly in PfMD3 knockout parasites, suggesting that PfMD3 impacts translation. In addition to RNA-binding, PfMD3 also interacts with proteins involved in mRNA processing and nuclear export that influence translational regulation. This study establishes PfMD3 as an RNA-binding protein that impacts lineage-specific translation and male gamete fertility.

molecular biology↗

bioRxiv2026

Dynamic regulation of long non-coding RNAs across asexual andgametocyte development in Plasmodium falciparum

Long non-coding RNAs (lncRNAs) are critical regulators of gene expression in eukaryotes. Short reads from Illumina sequencing, reverse transcriptase artefacts, and incomplete second-strand degradation in strand-specific cDNA libraries hamper genome-wide identification of lncRNAs, especially in gene-dense genomes such as Plasmodium. Here, we integrated long-read Oxford Nanopore Technology direct RNA sequencing, ribosome profiling, and single-cell transcriptomics to generate a robust and stage-specific characterization of P. falciparum lncRNAs. We generated comprehensive annotations of lncRNAs expressed in both asexual and sexual blood stages and confirmed their non-coding nature using ribosome profiling. Most lncRNAs showed pronounced stage-specific expression and appeared to be particularly abundant in mature gametocytes. Single-cell RNA sequencing revealed differential expression of many lncRNAs in female and male gametocytes, suggesting important roles in gametocytogenesis and transmission. Many lncRNAs are located antisense to protein-coding genes and are co-expressed with their sense mRNA, possibly from putative bidirectional promoters, while others overlap mRNA coding sequences or 3 untranslated regions and showed negatively correlated expression patterns. Overall, our study shows the prevalence of P. falciparum lncRNAs and highlights their possible roles in controlling the regulation of gene expression, particularly during gametocytogenesis.

genomics↗

bioRxiv2026

Double Mutations in Plasmodium falciparum Kelch13 drive resistance to next-generation artemisinin derivatives in malaria parasites

New antimalarial compounds are urgently required to overcome artemisinin partial resistance that has emerged in Asia and now Africa. Ozonides are promising next-generation artemisinins that offer the improved pharmacokinetic property of a prolonged in vivo half-life. To assess the potential for parasite resistance to ozonides in an artemisinin-resistant background, we subjected Cambodian Kelch13 (K13) mutant parasites to increasing artefenomel (OZ439) pressure up to in vivo physiological concentrations. Whole-genome sequencing identified a novel non-propeller K13 A212T mutation in OZ439-resistant parasites. Gene editing and drug susceptibility assays revealed that the K13 double mutation R539T+A212T is a determinant of OZ439 resistance. In extended parasite recovery assays, this resistance mechanism was associated with accelerated parasite recrudescence following OZ439 or OZ277 exposure. This phenotype was also observed in K13 C580Y+A212T double mutant parasites. Global metabolomic profiling revealed no changes in the levels of hemoglobin-derived peptides in OZ439-resistant parasites, suggesting that resistance is not associated with drug activation. Instead, double mutant parasites exhibited increased levels of metabolites linked to glutathione, nucleotide, and aspartate-glutamate metabolism, suggesting a higher capacity for redox regulation to tolerate drug-induced oxidative damage. Our findings demonstrate that ozonide resistance can emerge through a novel K13 mutation on the background of existing artemisinin-resistance k13 alleles.

microbiology↗

bioRxiv2025

Metabolomics reveals synergistic antimalarial drug pairing effects against Plasmodium falciparum in vitro

BackgroundThe development of new drugs against afflictions that disproportionately impact poorly resourced areas around the globe is an expensive endeavor. As cost-effective alternatives, strategic combinations of approved drugs can be used to enhance the efficacy against Plasmodium falciparum. Understanding the metabolic consequences of such combinations is essential for optimizing treatment strategies and delaying drug resistance. MethodsAn integrated metabolomic and pharmacological analysis was performed on P. falciparum exposed to chloroquine (CQ), pyrimethamine (PY), sulfadoxine (SD), and their combinations (SDPY and SDCQ). Dose{square}response assays were used to quantify drug potency, whereas untargeted metabolomic profiling was used to assess pathway-level perturbations associated with individual and combined treatments. ResultsDose{square}response assays confirmed the nanomolar potency of PY (IC = 12.5 nM) and CQ (IC = 11 nM) compared with the micromolar efficacy of SD (IC = 9.1 {micro}M), which is consistent with its role as a synergistic antifolate partner. Metabolomic profiling revealed that PY strongly disrupted folate-dependent pyrimidine biosynthesis, leading to deoxyuridine and dUMP accumulation, whereas SD caused milder perturbations, which was consistent with DHPS inhibition. CQ produced modest metabolic effects alone but markedly amplified antifolate-induced stress when combined with PY. Drug combinations generated metabolic responses that are distinct from those resulting from individual treatments. Across antifolate combinations, consistent trends included reduced amino acid pools, suppression of thiamine and glutathione metabolism, and enhanced PPP inhibition, leading to broad disruption of nucleotide, redox, and carbon metabolism. ConclusionsPyrimidine suppression has emerged as the central hallmark of antifolate-based therapy in P. falciparum. The distinct synergistic signatures observed with drug combinations support their potential to enhance efficacy and delay resistance. These findings provide a mechanistic foundation for guiding antimalarial combination policies, optimizing therapeutic regimens, and strengthening rational drug-design strategies.

systems biology↗

bioRxiv2025

Chromatin state dynamics during the Plasmodium falciparum intraerythrocytic development cycle

The interdependence of chromatin states and transcription factor (TF) binding in eukaryotic genomes is critical for the proper regulation of gene expression. In this study, we explore the connection between TFs and chromatin states in the human malaria parasite, Plasmodium falciparum, throughout its 48-hour asexual intraerythrocytic developmental cycle (IDC). Most P. falciparum genes are expressed in a periodic manner during the IDC, accompanied by dynamic shifts in histone modifications and chromatin accessibility. Leveraging genome-wide profiles of chromatin accessibility, histone modifications, and Heterochromatin Protein 1 (HP1) occupancy, we characterize chromatin state dynamics during the IDC. Our results indicate that several chromatin states remain stable throughout the lifecycle, while others are dynamic and are linked to gene activation or repression. We further characterize chromatin state dynamics at the genome-wide DNA binding sites for a selection of Plasmodium TFs, allowing us to group TFs according to their chromatin preferences. By correlating changes in chromatin accessibility, histone modifications, and TF binding, we provide a global overview of the chromatin state dynamics that coordinate P. falciparum asexual blood stage development.

genomics↗

bioRxiv2025

A single valine to leucine switch disrupts Plasmodium falciparum AP2-G DNA binding and reveals the role of GDV1 in ap2-g activation.

Sexual commitment in Plasmodium parasites is essential for malaria transmission, yet much remains unknown about the underlying signaling events initiating sexual conversion in a subpopulation of parasites. We discovered a single valine (V2163) to leucine (L2163) mutation in an Apetala 2 (AP2) transcription factor required for P. falciparum gametocytogenesis, ap2-g that abrogates sexual differentiation and confirmed this with forward and reverse mutation editing. Mutated AP2-G.L2163 does not bind the ap2-g consensus motif, GnGTAC, or stimulate AP2-G-dependent gene transcription including autoregulation. We then used AP2-G.L2163 parasite lines as tools to demonstrate the critical role of GDV1 in the initial activation of the silent ap2-g locus during the trophozoite to schizont transition in the absence of functional AP2-G and its autoregulation. Additionally, we show that AP2-G.V is required for MSRP1 expression, which can be used to distinguish early and late sexually committed schizonts. Together this work demonstrates that valine2163 in AP2-G plays a critical role in DNA binding, highlighting the functional importance of this specific region for malaria transmission as well as the critical role of GDV1 in the initial activation of ap2-g expression and induction of sexual differentiation. The reporter lines generated allow further study of signaling pathways or screening of factors regulating sexual commitment.

microbiology↗

bioRxiv2025

Degradation of ribosomal RNA during Plasmodium falciparum gametocytogenesis

The life cycle of Plasmodium falciparum is characterized by complex regulatory changes that allow adaptation of the parasites to different environmental conditions, which are especially pronounced during transmission between the mammalian host and the insect vector. Previous studies have shown that P. falciparum uses three types of ribosomal RNAs (rRNA A-, S1- and S2-types) at different stages of its life cycle. We used Oxford Nanopore Technologies (ONT) direct RNA sequencing to investigate the dynamics of rRNA usage throughout the parasites intraerythrocytic development, as well as in salivary gland sporozoites. Our study revealed a preponderance of A-type rRNAs during the intraerythrocytic cycle and gametocytogenesis, while S-type rRNAs slowly increase in abundance in mosquito stages starting three days post infection. Salivary gland sporozoites showed an even proportion of all rRNA types. By examining the length distributions of rRNA molecules, we detected an extensive and specific degradation of rRNAs during gametocytogenesis, starting in stage II gametocytes and continuing until the final stages of gametocyte development. We hypothesize that rRNA degradation may be linked to the global translational repression and metabolic quiescence described in stage V gametocytes, similar to mechanisms observed in bacterial and eukaryotic stress responses.

microbiology↗

bioRxiv2024

Hexosamine Biosynthesis Disruption Impairs GPI Production and Arrests Plasmodium falciparum Growth at Schizont Stages

UDP-N-acetylglucosamine (UDP-GlcNAc) is a crucial sugar nucleotide for glycan synthesis in eukaryotes. In the malaria parasite Plasmodium falciparum, UDP-GlcNAc is synthesized via the hexosamine biosynthetic pathway (HBP) and is essential for glycosylphosphatidylinositol (GPI) anchor production, the most prominent form of protein glycosylation in the parasite. In this study, we explore a conditional knockout of glucosamine-6-phosphate N-acetyltransferase (PfGNA1), a key HBP enzyme. PfGNA1 depletion led to significant disruptions in HBP metabolites, impairing GPI biosynthesis and causing mislocalization of the merozoite surface protein 1 (MSP1), the most abundant GPI-anchored protein in the parasite. Furthermore, parasites were arrested at the schizont stage, exhibiting severe segmentation defects and an incomplete rupture of the parasitophorous vacuole membrane (PVM), preventing egress from host red blood cells. Our findings demonstrate the critical role of HBP and GPI biosynthesis in P. falciparum asexual blood stage development and underscore the potential of targeting these pathways as a therapeutic strategy against malaria. Author SummaryMalaria remains a major cause of illness and death, particularly in sub-Saharan Africa, with increasing resistance to treatments highlighting the urgent need for new strategies. Malaria parasites rely on the hexosamine biosynthetic pathway to produce UDP-N-acetylglucosamine, an essential metabolite for glycosylphosphatidylinositol synthesis. Glycosylphosphatidylinositol molecules anchor vital proteins to the parasites surface and, as free glycolipids, serve as structural components of its membranes. Our study examined the effects of disrupting PfGNA1, a key enzyme in the hexosamine biosynthetic pathway, which is distinct from its human counterparts. Disruption of PfGNA1 blocked the production of glycosylphosphatidylinositol, leading to improper protein localization, developmental arrest, and failure of the parasites to mature or exit infected red blood cells. Our results underscore the central role of the hexosamine biosynthetic pathway and glycosylphosphatidylinositol biosynthesis, which are essential for parasite survival. This pathway represents a promising target for developing novel antimalarial therapies.

microbiology↗

bioRxiv2024

MULTIPLE, REDUNDANT CARBOXYLIC ACID TRANSPORTERS SUPPORT MITOCHONDRIAL METABOLISM IN PLASMODIUM FALCIPARUM

The mitochondrion of the deadliest human malaria parasite, Plasmodium falciparum, is an essential source of cellular acetyl-CoA during the asexual blood-stage of the parasite life cycle. Blocking mitochondrial acetyl-CoA synthesis leads to a hypoacetylated proteome and parasite death. We previously determined that mitochondrial acetyl-CoA is primarily synthesized from glucose-derived pyruvate by -ketoacid dehydrogenases. Here, we asked if inhibiting the import of glycolytic pyruvate across the mitochondrial inner membrane would affect acetyl-CoA production and, thus, could be a potential target for antimalarial drug development. We selected the two predicted mitochondrial pyruvate carrier proteins (PfMPC1 and PfMPC2) for genetic knockout and isotopic metabolite tracing via HPLC-MS metabolomic analysis. Surprisingly, we observed that asexual blood-stage parasites could survive the loss of either or both PfMPCs with only minor growth defects, despite a substantial reduction in the amount of glucose-derived isotopic labelling into acetyl-CoA. Furthermore, genetic deletion of two additional mitochondrial carboxylic acid transporters - DTC (di/tricarboxylic acid carrier) and YHM2 (a putative citrate/-ketoglutarate carrier protein) - only mildly affected asexual blood-stage replication, even in the context of PfMPC deficiency. Although we observed no added impact on the incorporation of glucose carbon into acetyl-CoA in these quadruple knockout mutants, we noted a large decrease in glutamine-derived label in tricarboxylic acid cycle metabolites, suggesting that DTC and YHM2 both import glutamine derivatives into the mitochondrion. Altogether, our results expose redundant routes used to fuel the blood-stage malaria parasite mitochondrion with imported carbon from two major sources - glucose and glutamine. SIGNIFICANCEThe mitochondrion of malaria parasites generates key molecules, such as acetyl-CoA, that are required for numerous cellular processes. To support mitochondrial biosynthetic pathways, the parasites must transport carbon sources into this organelle. By studying how the mitochondrion obtains pyruvate, a molecule derived from glucose, we have uncovered redundant carbon transport systems that ensure parasite survival in red blood cells. This metabolic redundancy poses a challenge for drug development, as it enables the parasite to adapt and survive by relying on alternative pathways when one is disrupted.

microbiology↗

bioRxiv2024

Structural characterization of the ACDC domain from ApiAP2 proteins of the malaria parasite

The Apicomplexan AP2 (ApiAP2) proteins are the best characterized family of DNA-binding proteins in the malaria parasite. Apart from the AP2 DNA-binding domain, there is little sequence similarity between ApiAP2 proteins and no other functional domains have been extensively characterized. One protein domain, which is present in a subset of the ApiAP2 proteins, is the conserved AP2-coincident domain mostly at the C-terminus (ACDC domain). Here we solved for the first time the crystal structure of the ACDC domain from two distinct Plasmodium falciparum ApiAP2 proteins and one orthologue from P. vivax, revealing a non-canonical four-helix bundle. Despite little sequence conservation between the ACDC domains from the two proteins, the structures are remarkably similar and do not resemble that of any other known protein domains. Due to their unique protein architecture and lack of homologues in the human genome, we performed in silico docking calculations against a library of known antimalarial compounds and we identified a small molecule that can potentially bind to any Apicomplexan ACDC domain within a pocket highly conserved amongst ApiAP2 proteins. Inhibitors based on this compound would disrupt the function of the ACDC domain and thus of the ApiAP2 proteins containing it, providing a new therapeutic window for targeting the malaria parasite and other Apicomplexans.

biochemistry↗

bioRxiv2023

Plasmodium falciparum MORC protein modulates gene expression through interaction with heterochromatin

Dynamic control of gene expression is critical for blood stage development of malaria parasites. Here, we used multi-omic analyses to investigate transcriptional regulation by the chromatin-associated microrchidia protein, MORC, during asexual blood stage development of the human malaria parasite Plasmodium falciparum. We show that PfMORC (PF3D7_1468100) interacts with a suite of nuclear proteins, including APETALA2 (AP2) transcription factors (PfAP2-G5, PfAP2-O5, PfAP2-I, PF3D7_0420300, PF3D7_0613800, PF3D7_1107800, and PF3D7_1239200), a DNA helicase DS60 (PF3D7_1227100), and other chromatin remodelers (PfCHD1 and PfEELM2). Transcriptomic analysis of PfMORCHA-glmS knockdown parasites revealed 163 differentially expressed genes belonging to hypervariable multigene families, along with upregulation of genes mostly involved in host cell invasion. In vivo genome-wide chromatin occupancy analysis during both trophozoite and schizont stages of development demonstrates that PfMORC is recruited to repressed, multigene families, including the var genes in subtelomeric chromosomal regions. Collectively, we find that PfMORC is found in chromatin complexes that play a role in the epigenetic control of asexual blood stage transcriptional regulation and chromatin organization.

microbiology↗

bioRxiv2023

DNA sequence context and the chromatin landscape differentiate sequence-specific transcription factor binding in the human malaria parasite, Plasmodium falciparum

Development of the human malaria parasite, Plasmodium falciparum, is regulated by a limited number of sequence-specific transcription factors (TFs). However, the mechanisms by which these TFs recognize genome-wide binding sites is still largely unknown. To address TF specificity, we investigated the binding of two TF subsets that either bind CACACA or GTGCAC DNA sequence motifs and further characterized PfAP2-G and PfAP2-EXP which bind unique DNA motifs (GTAC and TGCATGCA). We interrogated the impact of DNA sequence and chromatin context on P. falciparum TF binding by integrating high-throughput in vitro and in vivo binding assays, DNA shape predictions, epigenetic post-translational modifications, and chromatin accessibility. We determined that DNA sequence context minimally impacts binding site selection for CACACA-binding TFs, while chromatin accessibility, epigenetic patterns, co-factor recruitment, and dimerization contribute to differential binding. In contrast, GTGCAC-binding TFs prefer different DNA sequence context in addition to chromatin dynamics. Finally, we find that TFs that preferentially bind divergent DNA motifs may bind overlapping genomic regions in vivo due to low-affinity binding to other sequence motifs. Our results demonstrate that TF binding site selection relies on a combination of DNA sequence and chromatin features, thereby contributing to the complexity of P. falciparum gene regulatory mechanisms. Key PointsO_LISequence and chromatin context determine differential DNA-binding specificity of P. falciparum TFs C_LIO_LITFs with paralogous DNA-binding domains in P. falciparum are not functionally redundant C_LIO_LITFs with differing sequence-specificity can co-occupy sites through low-affinity DNA interactions C_LI

genomics↗

bioRxiv2022

Identification of Antimalarial Compounds that Inhibit Apicomplexan AP2 Proteins

Plasmodium parasites are reliant on the Apicomplexan AP2 (ApiAP2) transcription factor family to regulate gene expression programs. AP2 DNA binding domains have no homologs in the human or mosquito host genomes, making them potential antimalarial drug targets. Using an in-silico screen to dock thousands of small molecules into the crystal structure of the AP2-EXP (Pf3D7_1466400) AP2 domain (PDB:3IGM), we identified compounds that interact with this domain. Four compounds were found to compete for DNA binding with AP2-EXP and at least one additional ApiAP2 protein. Our top ApiAP2 competitor compound perturbs the transcriptome of P. falciparum trophozoites and results in a decrease in abundance of log2 fold change > 2 for 50% (46/93) of AP2-EXP target genes. Additionally, two ApiAP2 competitor compounds have multi-stage anti-Plasmodium activity against blood and mosquito stage parasites. In summary, we describe a novel set of antimalarial compounds that are targeted against the ApiAP2 family of proteins. These compounds may be used for future chemical genetic interrogation of ApiAP2 proteins or serve as starting points for a new class of antimalarial therapeutics. Author SummaryPlasmodium parasites are the causative agent of malaria, which resulted in over 600,000 deaths in 2021. Due to resistance arising for every antimalarial therapeutic deployed to date, new drug targets and druggable pathways must be explored. To address this concern, we used a molecular docking screen to predict competitors of DNA binding by the parasite specific family of Apicomplexan AP2 (ApiAP2) transcription factor proteins for testing in vitro and in vivo. We find that ApiAP2 competing compounds have antimalarial activity consistent with the disruption of gene regulation. This work will further our understanding of both the biological role and targetability of parasite transcriptional regulation.

molecular biology↗

bioRxiv2022

The mitochondrion of Plasmodium falciparum generates essential acetyl-CoA for protein acetylation

Coenzyme A (CoA) biosynthesis is an excellent target for antimalarial intervention. While most studies have focused on the use of CoA to produce acetyl-CoA in the apicoplast and the cytosol of malaria parasites, mitochondrial acetyl-CoA production is less well understood. In the current study, we performed metabolite labeling experiments to measure endogenous metabolites in Plasmodium falciparum lines with genetic deletions affecting mitochondrial dehydrogenase activity. Our results show that mitochondrial acetyl-CoA biosynthesis is essential for parasite growth and identify a catalytic redundancy between the two main ketoacid dehydrogenase enzymes, both of which are able to produce acetyl-CoA. The activity of these enzymes is dependent on the lipoate attachment enzyme LipL2, which is essential for parasite survival solely based on its role in supporting acetyl-CoA metabolism. We also find that acetyl-CoA produced in the mitochondrion is essential for the acetylation of histones and other proteins outside of the mitochondrion. Taken together, our results demonstrate that the mitochondrion is an essential de novo source of acetyl-CoA and is required for P. falciparum protein acetylation critical to parasite survival.

microbiology↗

bioRxiv2021

Preclinical characterization and target validation of the antimalarial pantothenamide MMV693183

Drug resistance and a dire lack of transmission-blocking antimalarials hamper malaria elimination. Here, we present the pantothenamide MMV693183 as a first-in-class acetyl-CoA synthetase (ACS) inhibitor to enter preclinical development. Our studies demonstrated attractive drug-like properties and in vivo efficacy in a humanized mouse model of Plasmodium falciparum infection. The compound showed exceptional in vitro activity against P. falciparum and P. vivax clinical isolates, and potently blocked P. falciparum transmission to Anopheles mosquitoes. Genetic and biochemical studies identified ACS as the target of the MMV693183-derived antimetabolite, CoA-MMV693183. MMV693183 was well adsorbed after oral administration in mice, rats and dogs. Pharmacokinetic - pharmacodynamic modelling predicted that a single 30 mg oral dose is sufficient to cure a malaria infection in humans. In conclusion, the ACS-targeting compound MMV693183 represents a promising addition to the portfolio of antimalarials in (pre)clinical development with a novel mode of action for the treatment of malaria and blocking transmission.

microbiology↗

bioRxiv2021

The PfAP2-HS transcription factor protects malaria parasites from febrile temperatures

Periodic fever is the most characteristic clinical feature of human malaria1-3, but how parasites survive febrile episodes is not known. While Plasmodium spp. genomes encode a full complement of chaperones4, they lack an ortholog of the conserved transcription factor HSF1, which in most eukaryotes activates the expression of key chaperones upon heat shock (HS)5-8. Here we identified PfAP2-HS, a transcription factor of the ApiAP2 family9-11, as the key regulator of the P. falciparum protective HS response. The PfAP2-HS-dependent HS response is largely restricted to rapid activation of hsp70-1, the predominant direct target of PfAP2-HS, and hsp90. Deletion of PfAP2-HS dramatically reduced HS survival and also resulted in severe growth defects at 37{degrees}C, but not at 35{degrees}C, and increased sensitivity to imbalances in protein homeostasis (proteostasis) produced by artemisinin, the current frontline antimalarial drug12,13. These results demonstrate that PfAP2-HS contributes to general maintenance of proteostasis and drives a rapid chaperone-based protective response against febrile temperatures. While several ApiAP2 transcription factors regulate life cycle transitions in malaria parasites11,14,15, PfAP2-HS is the first identified Plasmodium transcription factor that controls a protective response to a within-host environmental challenge.

microbiology↗

bioRxiv2020

The PfAP2-G2 transcription factor is a critical regulator of gametocyte maturation

Differentiation from asexual blood stages to sexual gametocytes is required for transmission of malaria parasites from the human to the mosquito host. Preventing gametocyte commitment and development would block parasite transmission, but the underlying molecular mechanisms behind these processes remain poorly understood. Here, we report that the ApiAP2 transcription factor, PfAP2-G2 (PF3D7_1408200) plays a critical role in the maturation of Plasmodium falciparum gametocytes. PfAP2-G2 binds to the promoters of a wide array of genes that are expressed at many stages of the parasite life cycle. Interestingly, we also find binding of PfAP2-G2 within the gene body of almost 3000 genes, which strongly correlates with the location of H3K36me3 and several other histone modifications as well as Heterochromatin Protein 1 (HP1), suggesting that occupancy of PfAP2-G2 in gene bodies may serve as an alternative regulatory mechanism. Disruption of pfap2-g2 does not impact asexual development, parasite multiplication rate, or commitment to sexual development but the majority of sexual parasites are unable to mature beyond stage III gametocytes. The absence of pfap2-g2 leads to overexpression of 28% of the genes bound by PfAP2-G2 and none of the PfAP2-g2 bound are downregulated, suggesting that it is a repressor. We also find that PfAP2-G2 interacts with chromatin remodeling proteins, a microrchidia (MORC) protein, and another ApiAP2 protein (PF3D7_1139300). Overall our data demonstrate that PfAP2-G2 is an important transcription factor that establishes an essential gametocyte maturation program in association with other chromatin-related proteins.

microbiology↗