{"gene":"PF3D7_1105000","run_date":"2026-09-08T12:51:28","timeline":{"discoveries":[{"year":2003,"finding":"The P. falciparum histone H4 gene is developmentally transcribed, with mRNA accumulating specifically in late-stage asexual (trophozoite/schizont) parasites, consistent with replication-coupled nucleosome assembly; the amino acid sequence is 100% conserved between P. falciparum, P. berghei, and P. yoelii.","method":"RT-PCR developmental transcription profiling; comparative sequence analysis","journal":"Parasitology research","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct transcription assay in parasites, replicated across three Plasmodium species at sequence level, single lab","pmids":["12739134"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""},{"species":"Plasmodium berghei","strain":""},{"species":"Plasmodium yoelii","strain":""}]},{"year":2006,"finding":"Mass spectrometry identified acetylation of lysines K5, K8, and K12 on histone H4 in P. falciparum; these marks were present at a relatively steady level throughout the erythrocytic cycle, while H4-K16 acetylation peaked in the mid-trophozoite stage. Methylation of lysines and arginines was also detected in the H4 N-terminus by MS.","method":"Mass spectrometry; immunoblotting with acetylation-specific antibodies","journal":"Gene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — two orthogonal methods (MS + antibody), single lab","pmids":["16410041"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2005,"finding":"Histone H4 hypoacetylation at telomere-proximal var gene loci is mediated by PfSir2; upon activation of a specific var gene, PfSir2 is removed from the promoter and histone acetylation (including H4) increases, linking H4 acetylation dynamics to mutually exclusive var gene expression.","method":"Chromatin immunoprecipitation (ChIP) with acetylation-specific antibodies; PfSir2 localization studies","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal ChIP with multiple antibodies, replicated across loci, mechanistic pathway placement established","pmids":["15820676"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2010,"finding":"Recombinant PfMYST HAT purified from P. falciparum parasites acetylates histone H4 in vitro at K5, K8, K12, and K16; overexpression of PfMYST in parasites altered global H4 acetylation levels and cell cycle progression.","method":"In vitro HAT assay with parasite-purified recombinant PfMYST; GFP-tagging and overexpression in parasites","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic assay with specific substrate identification plus cellular overexpression phenotype, single lab","pmids":["20807207"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2009,"finding":"Recombinant PfPRMT1 methylates histone H4 at Arg3 in vitro, forming monomethylarginine and asymmetric dimethylarginine; western blotting detected dynamic dimethylation of H4-Arg3 during parasite development, suggesting a role for this mark in transcription activation.","method":"In vitro methyltransferase assay; synthetic peptide MS for site mapping; immunoblotting with dimethyl-H4R3 antibody","journal":"The Biochemical journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with site-specific mapping by MS, corroborated by developmental western blotting, single lab","pmids":["19344311"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2008,"finding":"Methyl marks were detected on K20 of histone H4 in P. falciparum by site-specific methyl-lysine antibodies; two P. falciparum SET-domain proteins expressed in a eukaryotic system showed HKMT activity towards H4 in vitro.","method":"Immunoblotting with methyl-lysine-specific antibodies; in vitro HKMT assay in eukaryotic expression system","journal":"International journal for parasitology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — two orthogonal methods (antibody detection + in vitro enzymatic assay), single lab","pmids":["18299133"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2017,"finding":"H4K31 acetylation at gene promoters correlates with and may directly regulate gene expression in P. falciparum; monomethylated H4K31 is unexpectedly enriched at transcriptionally inactive pericentromeric heterochromatin in P. falciparum (a region devoid of H3K9me3 and HP1), establishing H4K31 modifications as determinants of genome organization.","method":"ChIP-seq; mass spectrometry identification of H4K31 modifications; comparative genomics with Toxoplasma gondii","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide ChIP-seq plus MS validation across two apicomplexan species, multiple orthogonal methods","pmids":["29101771"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2017,"finding":"Using a dominant-negative transgenic approach, H4K8 acetylation was identified as the rate-limiting regulatory step among H4 acetylations in P. falciparum; H4K8ac has a highly dynamic genome-wide distribution during the IDC, shifting from intergenic/upstream regions to protein-coding regions in association with transcriptional activity, and is present at var gene promoters in heterochromatin.","method":"Dominant-negative transgenic overexpression; ChIP-seq; HDAC inhibitor treatment","journal":"Epigenetics & chromatin","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic perturbation combined with genome-wide ChIP-seq and pharmacological validation, multiple orthogonal methods, single lab","pmids":["28830512"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"finding":"PfPRMT5 catalyzes symmetric dimethylation of histone H4 at R3 in vitro; PfPRMT5 disruption leads to depletion of active chromatin marks and down-regulation of invasion-related genes.","method":"In vitro methyltransferase assay; conditional knockdown; ChIP-seq; transcriptomics","journal":"Communications biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of H4R3 methylation activity plus genome-wide functional analysis via knockdown and ChIP-seq, single lab","pmids":["37349497"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2022,"finding":"The PfBDP1 bromodomain preferentially recognizes tetra-acetylated histone H4 (over other histone modifications) as shown by crystal structure, NMR spectroscopy, and ITC binding assays, indicating that histone H4 multi-acetylation serves as a binding platform for this P. falciparum epigenetic reader.","method":"Crystal structure of PfBDP1-BRD; solution NMR spectroscopy; isothermal titration calorimetry (ITC) with acetylated H4 peptides","journal":"International journal of biological macromolecules","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus NMR plus ITC, three orthogonal methods validating same interaction, single lab","pmids":["36328269"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2019,"finding":"Histone peptide pull-downs with acetylated H4 modifications revealed acetylation-level-dependent recruitment of the BDP1/BDP2 complex to histone H4; quantitative proteomics identified 14 putative reader proteins and their binding preferences for specific histone PTMs including H4 acetylations.","method":"Histone peptide pull-down; quantitative mass spectrometry; interaction proteomics","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — peptide pull-down with quantitative MS, single lab, establishes H4 acetylation as a reader recruitment signal","pmids":["31728527"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"finding":"Recombinant P. falciparum RbAp46/48 (PfRbAp46/48, PF3D7_0110700) directly interacts with histone H4 in vitro; PfRbAp46/48 localizes to the nucleus in all three asexual blood stages.","method":"Pull-down assay with purified recombinant PfRbAp46/48 and histone H4; nuclear fractionation/immunofluorescence","journal":"Parasitology research","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single pull-down assay, single lab, no reciprocal validation","pmids":["32363442"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2016,"finding":"Clinically approved HDAC inhibitors (belinostat, panobinostat, vorinostat, romidepsin) cause hyperacetylation of histone H4 in P. knowlesi parasites, confirming that HDAC enzymes in this Plasmodium species maintain H4 in a hypoacetylated state.","method":"In situ hyperacetylation assay (immunoblotting with anti-acetyl-H4 antibody) following HDAC inhibitor treatment","journal":"International journal for parasitology. Drugs and drug resistance","confidence":"Medium","confidence_rationale":"Tier 3 / Strong — single method but replicated across four structurally distinct inhibitors in the same study","pmids":["28107750"],"is_preprint":false,"attribution":[{"species":"Plasmodium knowlesi","strain":""}]},{"year":2021,"finding":"Cyclic tetrapeptide HDAC inhibitors cause hyperacetylation of histone H4 in P. falciparum parasites in situ, consistent with inhibition of one or more PfHDACs that act on H4.","method":"In situ hyperacetylation assay; PfHDAC1 catalytic activity inhibition assay","journal":"ACS infectious diseases","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single immunoblot readout for H4 hyperacetylation, pharmacological not genetic, single lab","pmids":["34491031"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]}],"current_model":"Plasmodium falciparum histone H4 is expressed in a replication-coupled manner peaking in late trophozoite/schizont stages; its N-terminal tail undergoes dynamic post-translational modifications including acetylation at K5, K8, K12, and K16 (written by PfMYST and erased by PfHDACs), arginine methylation at R3 (written by PfPRMT1 as an activating mark and by PfPRMT5 as symmetric dimethylation), and lysine methylation at K20; modifications within the globular domain (K31ac and K31me1) additionally shape genome organization and heterochromatin identity; tetra-acetylated H4 is recognized by the PfBDP1 bromodomain and acetylation-dependent reader complexes (BDP1/BDP2), while PfSir2-mediated H4 hypoacetylation propagates telomeric heterochromatin to silence var genes, and PfRbAp46/48 physically interacts with H4 as a chromatin-assembly chaperone."},"narrative":{"mechanistic_narrative":"Plasmodium falciparum histone H4 (PF3D7_1105000) is a core nucleosomal histone whose gene is transcribed in a replication-coupled manner, with mRNA peaking in late-stage asexual (trophozoite/schizont) parasites and an amino acid sequence that is fully conserved across P. falciparum, P. berghei, and P. yoelii [PMID:12739134]. Its N-terminal tail is a hub for dynamic post-translational modification that reads out into transcriptional control and genome organization. H4 is acetylated at K5, K8, K12, and K16, marks written in vitro by the acetyltransferase PfMYST and reversed by HDAC activity that maintains H4 in a hypoacetylated state [PMID:16410041, PMID:20807207, PMID:28107750]; among these, H4K8 acetylation is the rate-limiting acetylation and redistributes genome-wide from intergenic to coding regions in association with active transcription [PMID:28830512]. The tail is additionally methylated at R3—asymmetrically by PfPRMT1 and symmetrically by PfPRMT5, the latter promoting active chromatin and invasion-gene expression—and at K20 by SET-domain HKMTs [PMID:19344311, PMID:37349497, PMID:18299133]. Within the globular domain, K31 acetylation associates with active promoters while monomethyl-K31 demarcates pericentromeric heterochromatin, establishing H4K31 modifications as determinants of genome organization [PMID:29101771]. These modification states are decoded and propagated by dedicated machinery: PfSir2 enforces H4 hypoacetylation at telomere-proximal loci to silence var genes and is displaced upon var activation [PMID:15820676], while tetra-acetylated H4 is bound by the PfBDP1 bromodomain and recruits the BDP1/BDP2 reader complex in an acetylation-level-dependent manner [PMID:36328269, PMID:31728527]. The chromatin-assembly chaperone PfRbAp46/48 directly binds H4 in vitro [PMID:32363442].","teleology":[{"year":2003,"claim":"Establishing when H4 is made addressed whether its expression is coupled to DNA replication, framing H4 supply as part of the asexual cell cycle program.","evidence":"RT-PCR developmental transcription profiling and comparative sequence analysis across three Plasmodium species","pmids":["12739134"],"confidence":"Medium","gaps":["Does not measure protein-level abundance or nucleosome deposition timing","Does not address which chaperones deposit H4"],"attribution":[{"species":"Plasmodium falciparum","strain":""},{"species":"Plasmodium berghei","strain":""},{"species":"Plasmodium yoelii","strain":""}]},{"year":2005,"claim":"Linking H4 acetylation state to var gene silencing showed that reversible H4 hypoacetylation is a mechanism of mutually exclusive antigenic variation.","evidence":"ChIP with acetylation-specific antibodies and PfSir2 localization across var loci","pmids":["15820676"],"confidence":"High","gaps":["Does not resolve which specific H4 lysines PfSir2 targets","Does not establish how PfSir2 removal is triggered"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"claim":"Cataloguing H4 acetyl and methyl marks and their cell-cycle timing defined the modifiable residues and showed that individual marks differ in dynamics (steady K5/K8/K12 vs. trophozoite-peaking K16).","evidence":"Mass spectrometry and acetylation-specific immunoblotting across the erythrocytic cycle","pmids":["16410041"],"confidence":"Medium","gaps":["Does not assign enzymes to each mark","Does not map genomic distribution of the marks"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2008,"claim":"Detecting H4K20 methylation and SET-domain HKMT activity toward H4 extended the modification repertoire beyond acetylation to lysine methylation.","evidence":"Methyl-lysine-specific immunoblotting and in vitro HKMT assay from a eukaryotic expression system","pmids":["18299133"],"confidence":"Medium","gaps":["Does not identify which SET protein deposits K20me in vivo","Functional consequence of H4K20 methylation not established"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2009,"claim":"Identifying PfPRMT1 as the H4R3 methyltransferase assigned a writer to an arginine mark and connected it to transcriptional activation.","evidence":"In vitro methyltransferase assay with peptide MS site mapping and developmental immunoblotting","pmids":["19344311"],"confidence":"High","gaps":["In vivo requirement of PfPRMT1 for H4R3me not tested here","Downstream effectors reading H4R3me not identified"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2010,"claim":"Demonstrating that PfMYST acetylates H4 at K5/K8/K12/K16 assigned the writer for the H4 acetyl marks and linked H4 acetylation to cell-cycle progression.","evidence":"In vitro HAT assay with parasite-purified recombinant PfMYST plus GFP-tagged overexpression in parasites","pmids":["20807207"],"confidence":"High","gaps":["Does not distinguish PfMYST from other HATs at each site","Overexpression phenotype not resolved to specific H4 residues"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2016,"claim":"Showing that HDAC inhibitors hyperacetylate H4 confirmed that HDAC enzymes actively maintain H4 hypoacetylation, closing the write/erase loop for the acetyl marks (in P. knowlesi).","evidence":"In situ anti-acetyl-H4 immunoblotting after treatment with four clinical HDAC inhibitors","pmids":["28107750"],"confidence":"Medium","gaps":["Shown in P. knowlesi, not P. falciparum","Does not identify which specific HDAC erases which H4 site"],"attribution":[{"species":"Plasmodium knowlesi","strain":""}]},{"year":2017,"claim":"Genome-wide mapping identified H4K8ac as the rate-limiting acetylation and showed it redistributes with transcription, elevating H4K8ac to a principal regulatory mark of the intraerythrocytic cycle.","evidence":"Dominant-negative transgenic overexpression, ChIP-seq, and HDAC inhibitor treatment","pmids":["28830512"],"confidence":"High","gaps":["Mechanism coupling H4K8ac shifts to transcription machinery unresolved","Reader specific for H4K8ac not defined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2017,"claim":"Discovering globular-domain H4K31 modifications revealed that H4 shapes genome organization directly, with K31ac at active promoters and K31me1 marking an HP1/H3K9me3-independent pericentromeric heterochromatin.","evidence":"ChIP-seq, MS identification of H4K31 modifications, and comparative genomics with Toxoplasma gondii","pmids":["29101771"],"confidence":"High","gaps":["Writers/erasers of H4K31 modifications not identified","How K31me1 recruits or defines heterochromatin without HP1 unresolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2019,"claim":"Quantitative reader proteomics established that H4 acetylation level, not just presence, dictates recruitment of the BDP1/BDP2 complex and a set of other reader proteins.","evidence":"Histone peptide pull-down with quantitative mass spectrometry and interaction proteomics","pmids":["31728527"],"confidence":"Medium","gaps":["In vivo occupancy of readers on H4-marked chromatin not confirmed","Functional consequence of each reader binding not tested"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"claim":"Detecting a direct PfRbAp46/48–H4 interaction implicated a chromatin-assembly chaperone in H4 handling.","evidence":"Pull-down with purified recombinant PfRbAp46/48 and H4 plus nuclear localization by fractionation/IF","pmids":["32363442"],"confidence":"Low","gaps":["Single pull-down without reciprocal validation","In vivo role in H4 deposition not demonstrated","Binding interface not mapped"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2021,"claim":"Cyclic tetrapeptide HDAC inhibitors hyperacetylating H4 in P. falciparum reinforced that PfHDACs act on H4 in this species.","evidence":"In situ hyperacetylation immunoblot and PfHDAC1 catalytic inhibition assay","pmids":["34491031"],"confidence":"Low","gaps":["Single immunoblot readout, pharmacological not genetic","Does not map which H4 residues are deacetylated by which HDAC"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"}]},{"year":2022,"claim":"Structural characterization proved that the PfBDP1 bromodomain preferentially binds tetra-acetylated H4, defining H4 multi-acetylation as a discrete reader-binding platform.","evidence":"Crystal structure of PfBDP1-BRD, solution NMR, and ITC with acetylated H4 peptides","pmids":["36328269"],"confidence":"High","gaps":["Cellular consequence of BDP1 binding to tetra-acetyl H4 not tested here","Combinatorial specificity relative to single-acetyl marks not fully resolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2023,"claim":"Identifying PfPRMT5 as the H4R3 symmetric dimethyltransferase distinguished symmetric from asymmetric R3 methylation and tied it to active chromatin and invasion-gene regulation.","evidence":"In vitro methyltransferase assay, conditional knockdown, ChIP-seq, and transcriptomics","pmids":["37349497"],"confidence":"High","gaps":["Direct causal link between H4R3sme2 and specific invasion genes not isolated from other PfPRMT5 activities","Reader of symmetric H4R3me not identified"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"How the many H4 marks are integrated combinatorially to specify a given chromatin state, and which erasers act on the methyl and globular-domain marks, remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["Writers/erasers for H4K20me and H4K31 modifications not fully assigned","Combinatorial cross-talk among acetyl/methyl marks not mapped","In vivo requirement of most individual H4 marks for parasite viability untested"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["BDP1/BDP2 acetyl-H4 reader complex (H4 as bound ligand)"],"partners":["PFMYST (WRITER, H4K5/K8/K12/K16 ACETYLATION)","PFHDACS INCLUDING PFHDAC1 (ERASERS OF H4 ACETYLATION)","PFSIR2 (H4 HYPOACETYLATION AT TELOMERIC VAR LOCI)","PFPRMT1 (H4R3 ASYMMETRIC METHYLATION WRITER)","PFPRMT5 (H4R3 SYMMETRIC DIMETHYLATION WRITER)","SET-DOMAIN HKMTS (H4K20 METHYLATION)","PFBDP1 BROMODOMAIN (TETRA-ACETYL-H4 READER)","PFRBAP46/48 / PF3D7_0110700 (CHROMATIN-ASSEMBLY CHAPERONE, DIRECT H4 BINDING)"],"other_free_text":["H4 amino acid sequence 100% conserved across P. falciparum, P. berghei, and P. yoelii [#0].","H4K8 acetylation is the rate-limiting H4 acetylation with dynamic genome-wide redistribution during the intraerythrocytic cycle [#7].","Globular-domain H4K31ac marks active promoters while H4K31me1 marks HP1/H3K9me3-independent pericentromeric heterochromatin [#6].","HDAC-inhibitor-induced H4 hyperacetylation shown in P. falciparum [#13] and in P. knowlesi [#12]."]}},"prefetch_data":{},"corpus_meta":[{"pmid":"15820676","id":"PMC_15820676","title":"Telomeric heterochromatin propagation and histone acetylation control mutually exclusive expression of antigenic variation genes in malaria parasites.","date":"2005","source":"Cell","url":"https://pubmed.ncbi.nlm.nih.gov/15820676","citation_count":401,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"16410041","id":"PMC_16410041","title":"The malaria parasite Plasmodium falciparum histones: organization, expression, and 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Drugs and drug resistance","url":"https://pubmed.ncbi.nlm.nih.gov/40850017","citation_count":2,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["histone H4"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"41539273","id":"PMC_41539273","title":"Improved quantitative structure-activity relationship (QSAR) models to predict the activity of hydroxamic acids as HDAC inhibitors against malaria parasites.","date":"2025","source":"European journal of medicinal chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/41539273","citation_count":1,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["histone H4"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"31728527","id":"PMC_31728527","title":"Epigenetic reader complexes of the human malaria parasite, Plasmodium falciparum.","date":"2019","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/31728527","citation_count":48,"is_preprint":false,"source_species":["Pf"],"role":"citation_expansion","match_terms":["fwd_cite"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"30833341","id":"PMC_30833341","title":"Clipped histone H3 is integrated into nucleosomes of DNA replication genes in the human malaria parasite Plasmodium falciparum.","date":"2019","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/30833341","citation_count":11,"is_preprint":false,"source_species":["Pf"],"role":"citation_expansion","match_terms":["fwd_cite"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":14540,"output_tokens":3779,"usd":0.050153,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9573,"output_tokens":5410,"usd":0.091557,"stage2_stop_reason":"end_turn"},"total_usd":0.14171,"stage1_batch_id":"msgbatch_01DisAaTypwmC8QwdMBuePHo","stage2_batch_id":"msgbatch_0184zxZTqxiEfdYYfPMfkqjb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2003,\n      \"finding\": \"The P. falciparum histone H4 gene is developmentally transcribed, with mRNA accumulating specifically in late-stage asexual (trophozoite/schizont) parasites, consistent with replication-coupled nucleosome assembly; the amino acid sequence is 100% conserved between P. falciparum, P. berghei, and P. yoelii.\",\n      \"method\": \"RT-PCR developmental transcription profiling; comparative sequence analysis\",\n      \"journal\": \"Parasitology research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct transcription assay in parasites, replicated across three Plasmodium species at sequence level, single lab\",\n      \"pmids\": [\"12739134\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium berghei\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium yoelii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Mass spectrometry identified acetylation of lysines K5, K8, and K12 on histone H4 in P. falciparum; these marks were present at a relatively steady level throughout the erythrocytic cycle, while H4-K16 acetylation peaked in the mid-trophozoite stage. Methylation of lysines and arginines was also detected in the H4 N-terminus by MS.\",\n      \"method\": \"Mass spectrometry; immunoblotting with acetylation-specific antibodies\",\n      \"journal\": \"Gene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two orthogonal methods (MS + antibody), single lab\",\n      \"pmids\": [\"16410041\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Histone H4 hypoacetylation at telomere-proximal var gene loci is mediated by PfSir2; upon activation of a specific var gene, PfSir2 is removed from the promoter and histone acetylation (including H4) increases, linking H4 acetylation dynamics to mutually exclusive var gene expression.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP) with acetylation-specific antibodies; PfSir2 localization studies\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal ChIP with multiple antibodies, replicated across loci, mechanistic pathway placement established\",\n      \"pmids\": [\"15820676\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Recombinant PfMYST HAT purified from P. falciparum parasites acetylates histone H4 in vitro at K5, K8, K12, and K16; overexpression of PfMYST in parasites altered global H4 acetylation levels and cell cycle progression.\",\n      \"method\": \"In vitro HAT assay with parasite-purified recombinant PfMYST; GFP-tagging and overexpression in parasites\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic assay with specific substrate identification plus cellular overexpression phenotype, single lab\",\n      \"pmids\": [\"20807207\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Recombinant PfPRMT1 methylates histone H4 at Arg3 in vitro, forming monomethylarginine and asymmetric dimethylarginine; western blotting detected dynamic dimethylation of H4-Arg3 during parasite development, suggesting a role for this mark in transcription activation.\",\n      \"method\": \"In vitro methyltransferase assay; synthetic peptide MS for site mapping; immunoblotting with dimethyl-H4R3 antibody\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with site-specific mapping by MS, corroborated by developmental western blotting, single lab\",\n      \"pmids\": [\"19344311\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Methyl marks were detected on K20 of histone H4 in P. falciparum by site-specific methyl-lysine antibodies; two P. falciparum SET-domain proteins expressed in a eukaryotic system showed HKMT activity towards H4 in vitro.\",\n      \"method\": \"Immunoblotting with methyl-lysine-specific antibodies; in vitro HKMT assay in eukaryotic expression system\",\n      \"journal\": \"International journal for parasitology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two orthogonal methods (antibody detection + in vitro enzymatic assay), single lab\",\n      \"pmids\": [\"18299133\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"H4K31 acetylation at gene promoters correlates with and may directly regulate gene expression in P. falciparum; monomethylated H4K31 is unexpectedly enriched at transcriptionally inactive pericentromeric heterochromatin in P. falciparum (a region devoid of H3K9me3 and HP1), establishing H4K31 modifications as determinants of genome organization.\",\n      \"method\": \"ChIP-seq; mass spectrometry identification of H4K31 modifications; comparative genomics with Toxoplasma gondii\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide ChIP-seq plus MS validation across two apicomplexan species, multiple orthogonal methods\",\n      \"pmids\": [\"29101771\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Using a dominant-negative transgenic approach, H4K8 acetylation was identified as the rate-limiting regulatory step among H4 acetylations in P. falciparum; H4K8ac has a highly dynamic genome-wide distribution during the IDC, shifting from intergenic/upstream regions to protein-coding regions in association with transcriptional activity, and is present at var gene promoters in heterochromatin.\",\n      \"method\": \"Dominant-negative transgenic overexpression; ChIP-seq; HDAC inhibitor treatment\",\n      \"journal\": \"Epigenetics & chromatin\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic perturbation combined with genome-wide ChIP-seq and pharmacological validation, multiple orthogonal methods, single lab\",\n      \"pmids\": [\"28830512\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PfPRMT5 catalyzes symmetric dimethylation of histone H4 at R3 in vitro; PfPRMT5 disruption leads to depletion of active chromatin marks and down-regulation of invasion-related genes.\",\n      \"method\": \"In vitro methyltransferase assay; conditional knockdown; ChIP-seq; transcriptomics\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of H4R3 methylation activity plus genome-wide functional analysis via knockdown and ChIP-seq, single lab\",\n      \"pmids\": [\"37349497\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The PfBDP1 bromodomain preferentially recognizes tetra-acetylated histone H4 (over other histone modifications) as shown by crystal structure, NMR spectroscopy, and ITC binding assays, indicating that histone H4 multi-acetylation serves as a binding platform for this P. falciparum epigenetic reader.\",\n      \"method\": \"Crystal structure of PfBDP1-BRD; solution NMR spectroscopy; isothermal titration calorimetry (ITC) with acetylated H4 peptides\",\n      \"journal\": \"International journal of biological macromolecules\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus NMR plus ITC, three orthogonal methods validating same interaction, single lab\",\n      \"pmids\": [\"36328269\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Histone peptide pull-downs with acetylated H4 modifications revealed acetylation-level-dependent recruitment of the BDP1/BDP2 complex to histone H4; quantitative proteomics identified 14 putative reader proteins and their binding preferences for specific histone PTMs including H4 acetylations.\",\n      \"method\": \"Histone peptide pull-down; quantitative mass spectrometry; interaction proteomics\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — peptide pull-down with quantitative MS, single lab, establishes H4 acetylation as a reader recruitment signal\",\n      \"pmids\": [\"31728527\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Recombinant P. falciparum RbAp46/48 (PfRbAp46/48, PF3D7_0110700) directly interacts with histone H4 in vitro; PfRbAp46/48 localizes to the nucleus in all three asexual blood stages.\",\n      \"method\": \"Pull-down assay with purified recombinant PfRbAp46/48 and histone H4; nuclear fractionation/immunofluorescence\",\n      \"journal\": \"Parasitology research\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single pull-down assay, single lab, no reciprocal validation\",\n      \"pmids\": [\"32363442\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Clinically approved HDAC inhibitors (belinostat, panobinostat, vorinostat, romidepsin) cause hyperacetylation of histone H4 in P. knowlesi parasites, confirming that HDAC enzymes in this Plasmodium species maintain H4 in a hypoacetylated state.\",\n      \"method\": \"In situ hyperacetylation assay (immunoblotting with anti-acetyl-H4 antibody) following HDAC inhibitor treatment\",\n      \"journal\": \"International journal for parasitology. Drugs and drug resistance\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Strong — single method but replicated across four structurally distinct inhibitors in the same study\",\n      \"pmids\": [\"28107750\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium knowlesi\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Cyclic tetrapeptide HDAC inhibitors cause hyperacetylation of histone H4 in P. falciparum parasites in situ, consistent with inhibition of one or more PfHDACs that act on H4.\",\n      \"method\": \"In situ hyperacetylation assay; PfHDAC1 catalytic activity inhibition assay\",\n      \"journal\": \"ACS infectious diseases\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single immunoblot readout for H4 hyperacetylation, pharmacological not genetic, single lab\",\n      \"pmids\": [\"34491031\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}\n      ]\n    }\n  ],\n  \"current_model\": \"Plasmodium falciparum histone H4 is expressed in a replication-coupled manner peaking in late trophozoite/schizont stages; its N-terminal tail undergoes dynamic post-translational modifications including acetylation at K5, K8, K12, and K16 (written by PfMYST and erased by PfHDACs), arginine methylation at R3 (written by PfPRMT1 as an activating mark and by PfPRMT5 as symmetric dimethylation), and lysine methylation at K20; modifications within the globular domain (K31ac and K31me1) additionally shape genome organization and heterochromatin identity; tetra-acetylated H4 is recognized by the PfBDP1 bromodomain and acetylation-dependent reader complexes (BDP1/BDP2), while PfSir2-mediated H4 hypoacetylation propagates telomeric heterochromatin to silence var genes, and PfRbAp46/48 physically interacts with H4 as a chromatin-assembly chaperone.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"Plasmodium falciparum histone H4 (PF3D7_1105000) is a core nucleosomal histone whose gene is transcribed in a replication-coupled manner, with mRNA peaking in late-stage asexual (trophozoite/schizont) parasites and an amino acid sequence that is fully conserved across P. falciparum, P. berghei, and P. yoelii [#0]. Its N-terminal tail is a hub for dynamic post-translational modification that reads out into transcriptional control and genome organization. H4 is acetylated at K5, K8, K12, and K16, marks written in vitro by the acetyltransferase PfMYST and reversed by HDAC activity that maintains H4 in a hypoacetylated state [#1, #3, #12]; among these, H4K8 acetylation is the rate-limiting acetylation and redistributes genome-wide from intergenic to coding regions in association with active transcription [#7]. The tail is additionally methylated at R3\\u2014asymmetrically by PfPRMT1 and symmetrically by PfPRMT5, the latter promoting active chromatin and invasion-gene expression\\u2014and at K20 by SET-domain HKMTs [#4, #8, #5]. Within the globular domain, K31 acetylation associates with active promoters while monomethyl-K31 demarcates pericentromeric heterochromatin, establishing H4K31 modifications as determinants of genome organization [#6]. These modification states are decoded and propagated by dedicated machinery: PfSir2 enforces H4 hypoacetylation at telomere-proximal loci to silence var genes and is displaced upon var activation [#2], while tetra-acetylated H4 is bound by the PfBDP1 bromodomain and recruits the BDP1/BDP2 reader complex in an acetylation-level-dependent manner [#9, #10]. The chromatin-assembly chaperone PfRbAp46/48 directly binds H4 in vitro [#11].\",\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Establishing when H4 is made addressed whether its expression is coupled to DNA replication, framing H4 supply as part of the asexual cell cycle program.\",\n      \"evidence\": \"RT-PCR developmental transcription profiling and comparative sequence analysis across three Plasmodium species\",\n      \"pmids\": [\"12739134\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not measure protein-level abundance or nucleosome deposition timing\", \"Does not address which chaperones deposit H4\"],\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium berghei\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium yoelii\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Linking H4 acetylation state to var gene silencing showed that reversible H4 hypoacetylation is a mechanism of mutually exclusive antigenic variation.\",\n      \"evidence\": \"ChIP with acetylation-specific antibodies and PfSir2 localization across var loci\",\n      \"pmids\": [\"15820676\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve which specific H4 lysines PfSir2 targets\", \"Does not establish how PfSir2 removal is triggered\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Cataloguing H4 acetyl and methyl marks and their cell-cycle timing defined the modifiable residues and showed that individual marks differ in dynamics (steady K5/K8/K12 vs. trophozoite-peaking K16).\",\n      \"evidence\": \"Mass spectrometry and acetylation-specific immunoblotting across the erythrocytic cycle\",\n      \"pmids\": [\"16410041\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not assign enzymes to each mark\", \"Does not map genomic distribution of the marks\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Detecting H4K20 methylation and SET-domain HKMT activity toward H4 extended the modification repertoire beyond acetylation to lysine methylation.\",\n      \"evidence\": \"Methyl-lysine-specific immunoblotting and in vitro HKMT assay from a eukaryotic expression system\",\n      \"pmids\": [\"18299133\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not identify which SET protein deposits K20me in vivo\", \"Functional consequence of H4K20 methylation not established\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Identifying PfPRMT1 as the H4R3 methyltransferase assigned a writer to an arginine mark and connected it to transcriptional activation.\",\n      \"evidence\": \"In vitro methyltransferase assay with peptide MS site mapping and developmental immunoblotting\",\n      \"pmids\": [\"19344311\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo requirement of PfPRMT1 for H4R3me not tested here\", \"Downstream effectors reading H4R3me not identified\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Demonstrating that PfMYST acetylates H4 at K5/K8/K12/K16 assigned the writer for the H4 acetyl marks and linked H4 acetylation to cell-cycle progression.\",\n      \"evidence\": \"In vitro HAT assay with parasite-purified recombinant PfMYST plus GFP-tagged overexpression in parasites\",\n      \"pmids\": [\"20807207\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not distinguish PfMYST from other HATs at each site\", \"Overexpression phenotype not resolved to specific H4 residues\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Showing that HDAC inhibitors hyperacetylate H4 confirmed that HDAC enzymes actively maintain H4 hypoacetylation, closing the write/erase loop for the acetyl marks (in P. knowlesi).\",\n      \"evidence\": \"In situ anti-acetyl-H4 immunoblotting after treatment with four clinical HDAC inhibitors\",\n      \"pmids\": [\"28107750\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Shown in P. knowlesi, not P. falciparum\", \"Does not identify which specific HDAC erases which H4 site\"],\n      \"attribution\": [{\"species\": \"Plasmodium knowlesi\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Genome-wide mapping identified H4K8ac as the rate-limiting acetylation and showed it redistributes with transcription, elevating H4K8ac to a principal regulatory mark of the intraerythrocytic cycle.\",\n      \"evidence\": \"Dominant-negative transgenic overexpression, ChIP-seq, and HDAC inhibitor treatment\",\n      \"pmids\": [\"28830512\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism coupling H4K8ac shifts to transcription machinery unresolved\", \"Reader specific for H4K8ac not defined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Discovering globular-domain H4K31 modifications revealed that H4 shapes genome organization directly, with K31ac at active promoters and K31me1 marking an HP1/H3K9me3-independent pericentromeric heterochromatin.\",\n      \"evidence\": \"ChIP-seq, MS identification of H4K31 modifications, and comparative genomics with Toxoplasma gondii\",\n      \"pmids\": [\"29101771\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Writers/erasers of H4K31 modifications not identified\", \"How K31me1 recruits or defines heterochromatin without HP1 unresolved\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Quantitative reader proteomics established that H4 acetylation level, not just presence, dictates recruitment of the BDP1/BDP2 complex and a set of other reader proteins.\",\n      \"evidence\": \"Histone peptide pull-down with quantitative mass spectrometry and interaction proteomics\",\n      \"pmids\": [\"31728527\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vivo occupancy of readers on H4-marked chromatin not confirmed\", \"Functional consequence of each reader binding not tested\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Detecting a direct PfRbAp46/48\\u2013H4 interaction implicated a chromatin-assembly chaperone in H4 handling.\",\n      \"evidence\": \"Pull-down with purified recombinant PfRbAp46/48 and H4 plus nuclear localization by fractionation/IF\",\n      \"pmids\": [\"32363442\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single pull-down without reciprocal validation\", \"In vivo role in H4 deposition not demonstrated\", \"Binding interface not mapped\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Cyclic tetrapeptide HDAC inhibitors hyperacetylating H4 in P. falciparum reinforced that PfHDACs act on H4 in this species.\",\n      \"evidence\": \"In situ hyperacetylation immunoblot and PfHDAC1 catalytic inhibition assay\",\n      \"pmids\": [\"34491031\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single immunoblot readout, pharmacological not genetic\", \"Does not map which H4 residues are deacetylated by which HDAC\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"}]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Structural characterization proved that the PfBDP1 bromodomain preferentially binds tetra-acetylated H4, defining H4 multi-acetylation as a discrete reader-binding platform.\",\n      \"evidence\": \"Crystal structure of PfBDP1-BRD, solution NMR, and ITC with acetylated H4 peptides\",\n      \"pmids\": [\"36328269\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cellular consequence of BDP1 binding to tetra-acetyl H4 not tested here\", \"Combinatorial specificity relative to single-acetyl marks not fully resolved\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identifying PfPRMT5 as the H4R3 symmetric dimethyltransferase distinguished symmetric from asymmetric R3 methylation and tied it to active chromatin and invasion-gene regulation.\",\n      \"evidence\": \"In vitro methyltransferase assay, conditional knockdown, ChIP-seq, and transcriptomics\",\n      \"pmids\": [\"37349497\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct causal link between H4R3sme2 and specific invasion genes not isolated from other PfPRMT5 activities\", \"Reader of symmetric H4R3me not identified\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the many H4 marks are integrated combinatorially to specify a given chromatin state, and which erasers act on the methyl and globular-domain marks, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Writers/erasers for H4K20me and H4K31 modifications not fully assigned\", \"Combinatorial cross-talk among acetyl/methyl marks not mapped\", \"In vivo requirement of most individual H4 marks for parasite viability untested\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0000786\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0031492\", \"supporting_discovery_ids\": [9, 11]},\n      {\"term_id\": \"GO:0140033\", \"supporting_discovery_ids\": [9, 10]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0000785\", \"supporting_discovery_ids\": [2, 6, 7]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [11]},\n      {\"term_id\": \"GO:0005721\", \"supporting_discovery_ids\": [6]},\n      {\"term_id\": \"GO:0000781\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0006334\", \"supporting_discovery_ids\": [0, 11]},\n      {\"term_id\": \"GO:0006338\", \"supporting_discovery_ids\": [3, 7, 8]},\n      {\"term_id\": \"GO:0006342\", \"supporting_discovery_ids\": [2, 6]},\n      {\"term_id\": \"GO:0045814\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"GO:0006355\", \"supporting_discovery_ids\": [4, 8, 7]}\n    ],\n    \"complexes\": [\n      \"BDP1/BDP2 acetyl-H4 reader complex (H4 as bound ligand)\"\n    ],\n    \"partners\": [\n      \"PfMYST (writer, H4K5/K8/K12/K16 acetylation)\",\n      \"PfHDACs including PfHDAC1 (erasers of H4 acetylation)\",\n      \"PfSir2 (H4 hypoacetylation at telomeric var loci)\",\n      \"PfPRMT1 (H4R3 asymmetric methylation writer)\",\n      \"PfPRMT5 (H4R3 symmetric dimethylation writer)\",\n      \"SET-domain HKMTs (H4K20 methylation)\",\n      \"PfBDP1 bromodomain (tetra-acetyl-H4 reader)\",\n      \"PfRbAp46/48 / PF3D7_0110700 (chromatin-assembly chaperone, direct H4 binding)\"\n    ],\n    \"other_free_text\": [\n      \"H4 amino acid sequence 100% conserved across P. falciparum, P. berghei, and P. yoelii [#0].\",\n      \"H4K8 acetylation is the rate-limiting H4 acetylation with dynamic genome-wide redistribution during the intraerythrocytic cycle [#7].\",\n      \"Globular-domain H4K31ac marks active promoters while H4K31me1 marks HP1/H3K9me3-independent pericentromeric heterochromatin [#6].\",\n      \"HDAC-inhibitor-induced H4 hyperacetylation shown in P. falciparum [#13] and in P. knowlesi [#12].\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_1105000","name":null,"product":"histone H4","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_0941900","name":null,"product":"histone H4, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_0902600","name":null,"product":"histon H4, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"H4","evaluation":null}