{"gene":"H1-4","run_date":"2026-06-10T01:55:21","timeline":{"discoveries":[{"year":2005,"finding":"HP1 binds specifically to dimethylated Lys26 of histone H1.4 via its Chromo domain, and phosphorylation of the neighboring Ser27 prevents this binding, establishing a 'phospho-switch' mechanism.","method":"In vitro binding assays with modified H1.4 peptides and HP1 Chromo domain; mutagenesis of S27","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct in vitro binding with domain mapping and mutagenesis, replicated in later studies confirming the same mechanism","pmids":["16127177"],"is_preprint":false},{"year":2011,"finding":"Aurora B kinase phosphorylates H1.4 specifically at Ser27 in a cell-cycle-regulated manner peaking at metaphase; adjacent K26 dimethylation modulates Aurora B activity toward S27; H1.4 is the only somatic linker histone variant targeted by Aurora B; S27 phosphorylation increases H1.4 mobility and reduces chromatin binding in mitosis.","method":"In vitro kinase assays; in vivo phosphorylation analysis by immunofluorescence and immunoblot; FRAP on H1.4 mutants; cell-cycle fractionation","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — in vitro kinase assay plus FRAP in vivo, multiple orthogonal methods in single study","pmids":["21511733"],"is_preprint":false},{"year":2011,"finding":"Protein kinase A (PKA) phosphorylates H1.4 at Ser35; this phosphorylation causes H1.4 to dissociate from mitotic chromatin, and an H1.4-S35A mutant cannot rescue mitotic defects after H1.4 depletion; inhibition of PKA increases mitotic chromatin compaction in an H1.4-dependent manner.","method":"Mass spectrometry identification of phosphorylation site; in vitro kinase assay; mutant rescue experiments; PKA inhibitor treatment with chromatin compaction readout","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — in vitro kinase assay plus mutant rescue plus pharmacological inhibition, multiple orthogonal methods","pmids":["21852232"],"is_preprint":false},{"year":2012,"finding":"GCN5 acetylates H1.4 at Lys34 (H1.4K34ac); this mark is enriched at promoters of active genes, increases H1.4 mobility (measured by FRAP), and recruits a general transcription factor to stimulate transcription.","method":"In vitro acetylation assay with GCN5; ChIP-seq for H1.4K34ac; FRAP of H1.4 mutants; transcription factor recruitment assays","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 1-2 / Moderate — in vitro enzymatic assay combined with ChIP, FRAP, and functional transcription data in a single study","pmids":["22465951"],"is_preprint":false},{"year":2013,"finding":"SET7/9 methylates H1.4 at multiple lysine residues within KAK motifs in the C-terminal domain (K121, K129, K159, K171, K177, K192); ARTD1/PARP1-mediated poly(ADP-ribosyl)ation of H3 enables subsequent SET7/9-dependent methylation of H1.4; H1.4 and H3 compete for SET7/9 methylation.","method":"In vitro methylation assays with isolated histones; poly(ADP-ribosyl)ation assays; mass spectrometry identification of methylation sites","journal":"Epigenetics & chromatin","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro biochemical reconstitution but single lab, no in vivo validation reported","pmids":["23289424"],"is_preprint":false},{"year":2008,"finding":"Histone H1e (H1.4) physically interacts with the small hepatitis delta antigen (SHDAg) via H1.4's central globular domain; this interaction requires SHDAg oligomerization; N- or C-terminal deletion mutants of H1e inhibit HDV replication, which is rescued by wild-type H1e.","method":"Tandem affinity purification/mass spectrometry; domain-mapping by deletion mutagenesis; HDV replication rescue assays","journal":"Virology","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — TAP-MS identification plus domain mapping and functional rescue, single lab","pmids":["18314153"],"is_preprint":false},{"year":2019,"finding":"Frameshift mutations in the C-terminal tail of H1.4 produce stable nuclear proteins that bind chromatin, disrupt proper DNA compaction, are associated with a specific methylation pattern, reduce cell proliferation and S-phase entry, and accelerate cellular senescence.","method":"Expression of mutant H1.4 in cells; nuclear fractionation; DAPI staining for chromatin compaction; BrdU/flow cytometry for cell cycle; senescence assays; methylome analysis","journal":"American journal of human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple cellular assays with defined molecular readouts, single lab","pmids":["31447100"],"is_preprint":false},{"year":2020,"finding":"Loss of H1.4 (and H1.2) in human PLB-985 cells induces an eosinophil-like transcriptional program and negatively regulates neutrophil lineage differentiation; this subtype-specific effect is confirmed in murine bone marrow stem cells.","method":"Genome-wide CRISPR/Cas9 screen; systematic disruption of individual H1 subtypes; transcriptional profiling; murine bone marrow differentiation assays","journal":"eLife","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with transcriptomic readout in two systems (human cell line and murine primary cells), single lab","pmids":["32391789"],"is_preprint":false},{"year":2020,"finding":"Phosphorylated H1.4 at serine 187 (pS187-H1.4) remains associated with active promoters genome-wide (enriched at transcription start sites of estrogen-activated genes), stably interacts with RNA Polymerase II, and is proposed to be phosphorylated by CDK9 as an early event in gene activation.","method":"ChIP-seq with pS187-H1.4-specific antibodies in MCF7 cells; co-immunoprecipitation with RNAPII; estradiol induction system","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genome-wide ChIP-seq with specific antibodies plus co-IP with RNAPII, single lab","pmids":["33238524"],"is_preprint":false},{"year":2022,"finding":"Exogenous expression of H1.4 C-terminal frameshift mutant (H1.4 CFT) in rat hippocampal neurons alters expression of ~400 genes (downregulating synaptic communication and neuropeptide signaling genes) and reduces neuronal activity as measured by multielectrode arrays.","method":"Exogenous expression of WT or mutant H1.4 in rat hippocampal neurons; genome-wide transcriptome analysis (RNA-seq); multielectrode array recording","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — transcriptome plus functional electrophysiology, single lab","pmids":["34788807"],"is_preprint":false},{"year":2026,"finding":"A Rahman syndrome frameshift mutation in H1.4's C-terminal domain causes more extended/flexible nucleosome array conformation, enhanced linker DNA accessibility, inability to form compact stacked nucleosome structures, reduced capacity for liquid-liquid and liquid-solid phase separation, and increased H1.4 nuclear mobility; molecular dynamics simulations show the mutated CTD interacts with a shorter linker DNA segment.","method":"In vitro reconstitution of hexanucleosomal arrays; cryo-EM/structural analysis; FRET measurements; molecular dynamics simulations; FRAP in cells; micrococcal nuclease digestion","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — reconstitution with structural analysis, MD simulations, FRET, and FRAP, multiple orthogonal methods in single rigorous study","pmids":["42173878"],"is_preprint":false},{"year":2025,"finding":"H1.4 ablation causes robust changes in nascent transcription and gene expression; loss of H1.4 alters chromatin accessibility at enhancer-sized extragenic and intronic regions with concordant changes in H3K27ac/H3K4me1; sites losing accessibility are enriched for AP-1 motifs, placing H1.4 as a regulator of AP-1-directed enhancers.","method":"H1.4 knockout; nascent transcription assays (PRO-seq or equivalent); ATAC-seq; ChIP for H3K27ac and H3K4me1; motif analysis","journal":"iScience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO with ATAC-seq and histone modification ChIP, multiple orthogonal methods, single lab","pmids":["41054521"],"is_preprint":false},{"year":1994,"finding":"H1e (H1.4) binds preferentially and cooperatively to GC-rich DNA; a 25-residue C-terminal domain peptide retains GC-rich sequence preference but not cooperativity.","method":"DNA melting analysis; gel-mobility-shift assay; synthetic peptide binding experiments","journal":"Biochemistry","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro biochemical assays with purified protein and synthetic peptides, single lab","pmids":["8286360"],"is_preprint":false},{"year":1996,"finding":"H1e (H1.4) is the only somatic linker histone variant that forms H1-H1 polymers when bound to oligonucleosomal DNA; poly(ADP-ribosyl)ation of H1e reduces polymer size without displacing it from linker regions.","method":"Cross-linking analysis; reverse-phase HPLC purification of H1 variants; methyl-accepting ability assay in native nuclei","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — biochemical reconstitution with purified variants and cross-linking, single lab","pmids":["8687390"],"is_preprint":false},{"year":1995,"finding":"H1e (H1.4) variants specifically inhibit in vitro enzymatic DNA methylation and preferentially bind unmethylated CpG-rich DNA compared to H1c; the C-terminal domain mediates preferential binding to CpG-rich sequences.","method":"In vitro DNA methylation assay; gel retardation; Southwestern blot with CpG-rich oligonucleotides; competition binding assays","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro enzymatic and binding assays, single lab, replicated in companion 1996 paper","pmids":["7848272"],"is_preprint":false},{"year":2024,"finding":"CYP1B1 physically associates with linker histone H1.4 (identified by LC-MS); downregulation of H1.4 is associated with increased chromatin accessibility and higher cell viability after PARP inhibitor treatment in resistant ovarian cancer cells.","method":"LC-MS proteomics for CYP1B1 interactors; ATAC-seq; H1.4 knockdown with cell viability readout; micrococcal nuclease digestion","journal":"Drug resistance updates","confidence":"Low","confidence_rationale":"Tier 3 / Weak — LC-MS identification of interaction without reciprocal validation; functional link is correlative, single lab","pmids":["39395328"],"is_preprint":false},{"year":2025,"finding":"Simultaneous knockdown of H1.2, H1.3, H1.4, and H1.5 in K562 cells reverses silencing of unintegrated HIV-1 DNA, resulting in increased viral expression; this effect is specific to HIV-1 and not observed for MLV unintegrated DNA.","method":"siRNA knockdown of four H1 variants; RT-qPCR/reporter assay for unintegrated HIV-1 and MLV expression","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — combined H1 subtype knockdown does not isolate H1.4-specific contribution; preprint, single method","pmids":[],"is_preprint":true},{"year":2025,"finding":"SETDB1 binds to SUMOylated histone H1.2 and H1.4; SUMOylated H1.4 colocalizes with H3K9me3 at repetitive regions of the genome.","method":"PLAMseq (TurboID proximity labeling coupled to mass spectrometry and sequencing); SUMOylation mapping","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — proximity labeling approach without reciprocal validation; preprint, single method","pmids":[],"is_preprint":true},{"year":2002,"finding":"The N-terminal domain peptide of histone H1e (residues 15-36) adopts two amphipathic alpha-helices separated by a flexible Gly-Gly motif under helix-stabilizing conditions; the Gly-Gly motif allows a wide range of relative orientations potentially facilitating DNA backbone tracking or simultaneous binding of two DNA segments.","method":"CD spectroscopy; 1H-NMR structure calculation in TFE solution","journal":"Protein science","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — NMR structure determination with CD validation, but performed on isolated peptide in non-physiological conditions, single lab","pmids":["11790831"],"is_preprint":false}],"current_model":"Histone H1.4 is a linker histone that compacts chromatin by binding linker DNA (preferentially GC-rich sequences) and forming H1-H1 polymers on nucleosomal arrays; its functions are regulated by an array of post-translational modifications—including Aurora B-mediated S27 phosphorylation (which evicts HP1 bound to dimethyl-K26 and increases H1.4 mobility at mitosis), PKA-mediated S35 phosphorylation (which dissociates H1.4 from mitotic chromatin), CDK9/S187 phosphorylation (which keeps H1.4 at active promoters and promotes interaction with RNA Pol II), GCN5-mediated K34 acetylation (which increases mobility and recruits general transcription factors to active genes), and SET7/9-mediated C-terminal lysine methylation (modulated by ARTD1/PARP1 activity); disease-associated frameshift mutations in the C-terminal domain disrupt chromatin compaction, phase separation, and gene regulation, causing the neurodevelopmental Rahman syndrome."},"narrative":{"mechanistic_narrative":"Histone H1.4 is a somatic linker histone that compacts chromatin by binding linker DNA—preferentially GC-rich and unmethylated CpG-rich sequences through its C-terminal domain—and is the only somatic H1 variant that forms H1-H1 polymers on oligonucleosomal arrays [PMID:8286360, PMID:8687390, PMID:7848272]. Functional reconstitution shows that H1.4 drives formation of compact, stacked nucleosome structures and supports liquid-liquid/liquid-solid phase separation, with its C-terminal tail engaging linker DNA to limit accessibility [PMID:42173878]. Its chromatin-binding behavior is tuned by a dense array of post-translational modifications: Aurora B phosphorylates Ser27 in a metaphase-peaking manner to increase H1.4 mobility and evict HP1, which otherwise reads dimethyl-Lys26 via a phospho-switch [PMID:16127177, PMID:21511733]; PKA phosphorylates Ser35 to dissociate H1.4 from mitotic chromatin [PMID:21852232]; GCN5 acetylates Lys34 at active promoters to increase mobility and recruit general transcription factors [PMID:22465951]; CDK9-dependent Ser187 phosphorylation keeps H1.4 at active transcription start sites in stable association with RNA Polymerase II [PMID:33238524]; and SET7/9 methylates C-terminal KAK-motif lysines in a manner gated by PARP1/ARTD1 activity [PMID:23289424]. Consistent with these roles, H1.4 loss reprograms nascent transcription and chromatin accessibility at AP-1-bound enhancers and biases hematopoietic differentiation [PMID:41054521, PMID:32391789]. C-terminal frameshift mutations that produce stable but functionally aberrant nuclear H1.4 cause the neurodevelopmental Rahman syndrome; these mutants fail to compact chromatin or phase separate, reduce proliferation and accelerate senescence, and in neurons downregulate synaptic-signaling genes and reduce neuronal activity [PMID:31447100, PMID:34788807, PMID:42173878].","teleology":[{"year":1994,"claim":"Established the intrinsic DNA-binding preference of H1.4, showing it recognizes GC-rich linker DNA cooperatively through its C-terminal domain rather than binding DNA indiscriminately.","evidence":"DNA melting, gel-shift, and synthetic C-terminal peptide binding assays","pmids":["8286360"],"confidence":"Medium","gaps":["No structural basis for sequence preference","Cooperativity determinant outside the tested 25-residue peptide not mapped"]},{"year":1995,"claim":"Linked H1.4 DNA-binding specificity to epigenetic state by showing it preferentially binds unmethylated CpG-rich DNA and inhibits DNA methylation in vitro, distinguishing it functionally from H1c.","evidence":"In vitro methylation, gel retardation, and Southwestern assays with CpG-rich oligonucleotides","pmids":["7848272"],"confidence":"Medium","gaps":["In vitro only; no cellular demonstration of methylation regulation","Mechanism of methyltransferase inhibition unresolved"]},{"year":1996,"claim":"Identified H1.4 as the unique somatic variant capable of self-polymerization on nucleosomal arrays, implicating it in higher-order chromatin organization, and showed poly(ADP-ribosyl)ation modulates polymer size.","evidence":"Cross-linking, HPLC variant purification, and methyl-accepting assays in native nuclei","pmids":["8687390"],"confidence":"Medium","gaps":["Functional consequence of polymer formation for chromatin compaction not directly tested in vivo","Polymerization interface not mapped"]},{"year":2002,"claim":"Provided structural rationale for linker DNA engagement by showing the H1e N-terminal peptide forms two amphipathic helices joined by a flexible Gly-Gly hinge that could allow DNA tracking or dual-segment binding.","evidence":"CD spectroscopy and 1H-NMR structure determination in TFE","pmids":["11790831"],"confidence":"Medium","gaps":["Isolated peptide in non-physiological solvent","No structure on nucleosomal substrate"]},{"year":2005,"claim":"Defined a phospho-methyl switch on the H1.4 N-terminal tail in which HP1 reads dimethyl-Lys26 and adjacent Ser27 phosphorylation blocks it, coupling H1.4 modification to heterochromatin reader recruitment.","evidence":"In vitro peptide-binding assays with the HP1 Chromo domain and S27 mutagenesis","pmids":["16127177"],"confidence":"High","gaps":["Kinase responsible for S27 not identified in this study","In vivo dynamics of the switch not measured here"]},{"year":2011,"claim":"Identified the mitotic kinases acting on the H1.4 tail: Aurora B phosphorylates S27 (peaking at metaphase, modulated by K26 methylation) and PKA phosphorylates S35, both reducing H1.4 chromatin binding, connecting the phospho-switch to mitotic chromatin dynamics.","evidence":"In vitro kinase assays, immunofluorescence/immunoblot, FRAP, mutant rescue, and PKA inhibition with compaction readouts","pmids":["21511733","21852232"],"confidence":"High","gaps":["Phosphatases reversing these marks not identified","Interplay between S27 and S35 phosphorylation not resolved"]},{"year":2012,"claim":"Connected H1.4 to active transcription by showing GCN5-mediated K34 acetylation marks active promoters, increases H1.4 mobility, and recruits a general transcription factor, recasting linker histone modification as activating rather than purely repressive.","evidence":"In vitro acetylation, ChIP-seq for H1.4K34ac, FRAP, and transcription factor recruitment assays","pmids":["22465951"],"confidence":"High","gaps":["Identity of the recruited GTF interaction surface not fully defined","Whether K34ac and other marks co-occur on the same molecule unknown"]},{"year":2013,"claim":"Mapped SET7/9 methylation of multiple C-terminal KAK-motif lysines on H1.4 and showed PARP1/ARTD1 ribosylation of H3 gates this methylation, linking H1.4 modification to a broader PARP-dependent regulatory network.","evidence":"In vitro methylation and poly(ADP-ribosyl)ation assays with mass spectrometry site mapping","pmids":["23289424"],"confidence":"Medium","gaps":["No in vivo validation of the methylation sites","Functional consequence of C-terminal methylation untested"]},{"year":2008,"claim":"Demonstrated a host-pathogen role in which H1.4's globular domain binds oligomerized small hepatitis delta antigen and supports HDV replication, the first non-chromatin functional partner identified.","evidence":"TAP-MS, deletion domain mapping, and HDV replication rescue assays","pmids":["18314153"],"confidence":"Medium","gaps":["Mechanism by which H1.4 promotes HDV replication unclear","Single lab, no reciprocal in vivo validation"]},{"year":2019,"claim":"Established H1.4 C-terminal frameshift mutations as causative of a disease state, showing the stable mutant proteins bind chromatin but disrupt compaction, alter methylation, reduce proliferation, and accelerate senescence.","evidence":"Expression of mutant H1.4 with fractionation, DAPI compaction, cell-cycle, senescence, and methylome assays","pmids":["31447100"],"confidence":"Medium","gaps":["Mechanistic link from compaction defect to senescence not fully resolved","Single lab"]},{"year":2020,"claim":"Revealed two distinct in vivo roles: pS187-H1.4 (CDK9-dependent) marks active promoters and binds RNA Pol II, while loss of H1.4 biases myeloid differentiation toward an eosinophil-like program, showing subtype-specific regulatory function.","evidence":"pS187 ChIP-seq and RNAPII co-IP in MCF7; genome-wide CRISPR screen and transcriptional/differentiation profiling in PLB-985 and murine bone marrow","pmids":["33238524","32391789"],"confidence":"Medium","gaps":["Direct demonstration that CDK9 phosphorylates S187 not shown","How H1.4 loss selects lineage outcome mechanistically unclear"]},{"year":2022,"claim":"Connected the H1.4 C-terminal mutant to the neuronal phenotype of disease by showing it dysregulates ~400 genes, downregulating synaptic and neuropeptide signaling, and reduces neuronal electrical activity.","evidence":"Exogenous WT/mutant H1.4 expression in rat hippocampal neurons with RNA-seq and multielectrode array recording","pmids":["34788807"],"confidence":"Medium","gaps":["Direct chromatin mechanism at affected neuronal genes not mapped","Exogenous overexpression may not model endogenous dosage"]},{"year":2025,"claim":"Placed H1.4 as a regulator of AP-1-directed enhancers, showing its loss alters chromatin accessibility and enhancer histone marks at extragenic/intronic sites with concordant nascent transcription changes.","evidence":"H1.4 knockout with PRO-seq, ATAC-seq, H3K27ac/H3K4me1 ChIP, and motif analysis","pmids":["41054521"],"confidence":"Medium","gaps":["Whether H1.4 directly opposes AP-1 binding or acts indirectly unclear","Single lab"]},{"year":2026,"claim":"Provided the structural and biophysical mechanism of Rahman syndrome, showing a C-terminal frameshift mutant produces extended nucleosome arrays, increased linker DNA accessibility, failed compaction, impaired phase separation, and higher nuclear mobility.","evidence":"In vitro hexanucleosome reconstitution, cryo-EM/structural analysis, FRET, molecular dynamics, FRAP, and MNase digestion","pmids":["42173878"],"confidence":"High","gaps":["How phase-separation loss translates to specific gene dysregulation not directly linked","Effect of disease mutant on PTM-mediated regulation untested"]},{"year":null,"claim":"It remains unknown how the combinatorial array of H1.4 post-translational modifications is integrated on individual molecules to switch between compaction, phase separation, and active-promoter functions, and how disease mutations perturb this integrated code in specific cell types.","evidence":"No single study in the corpus reconstitutes the full modification network on defined chromatin","pmids":[],"confidence":"Low","gaps":["No combinatorial PTM analysis on single molecules","Phosphatases/demethylases for H1.4 marks unidentified","Cell-type-specific rules linking H1.4 dosage to gene programs undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[12,14,10]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[10,13]}],"localization":[{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[1,6,10]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[6,8]}],"pathway":[{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[3,10,11]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[3,8,11]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[1,2]}],"complexes":[],"partners":["HP1","AURKB","PRKACA","GCN5","SETD7","RNA POL II","PARP1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P10412","full_name":"Histone H1.4","aliases":["Histone H1b","Histone H1s-4"],"length_aa":219,"mass_kda":21.9,"function":"Histone H1 protein binds to linker DNA between nucleosomes forming the macromolecular structure known as the chromatin fiber (PubMed:35581345, PubMed:40240600). Histones H1 are necessary for the condensation of nucleosome chains into higher-order structured fibers and promote formation of the H3K27me3 mark by the PRC2/EED-EZH2 complex (PubMed:35581345, PubMed:40240600, PubMed:40516528). Ability to associate with nucleosomes and compact chromatin depends on linker DNA length and trajectory (PubMed:35581345). Also acts as a regulator of individual gene transcription through chromatin remodeling, nucleosome spacing and DNA methylation (PubMed:40240600)","subcellular_location":"Nucleus; Chromosome","url":"https://www.uniprot.org/uniprotkb/P10412/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/H1-4","classification":"Not Classified","n_dependent_lines":163,"n_total_lines":1208,"dependency_fraction":0.13493377483443708},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"HIST2H2BE","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/H1-4","total_profiled":1310},"omim":[{"mim_id":"617910","title":"LSM11, U7 SMALL NUCLEAR RNA-ASSOCIATED PROTEIN; LSM11","url":"https://www.omim.org/entry/617910"},{"mim_id":"617876","title":"RNA, U7 SMALL NUCLEAR 1; RNU7-1","url":"https://www.omim.org/entry/617876"},{"mim_id":"617537","title":"RAHMAN SYNDROME; RMNS","url":"https://www.omim.org/entry/617537"},{"mim_id":"609693","title":"VON WILLEBRAND FACTOR A DOMAIN-CONTAINING PROTEIN 7; VWA7","url":"https://www.omim.org/entry/609693"},{"mim_id":"142712","title":"HISTONE GENE CLUSTER 1, H1 HISTONE FAMILY, MEMBER T; HIST1H1T","url":"https://www.omim.org/entry/142712"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nuclear speckles","reliability":"Supported"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"bone marrow","ntpm":49.4}],"url":"https://www.proteinatlas.org/search/H1-4"},"hgnc":{"alias_symbol":["H1.4","H1e","H1s-4"],"prev_symbol":["H1F4","HIST1H1E"]},"alphafold":{"accession":"P10412","domains":[{"cath_id":"1.10.10","chopping":"58-109","consensus_level":"medium","plddt":94.7923,"start":58,"end":109}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P10412","model_url":"https://alphafold.ebi.ac.uk/files/AF-P10412-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P10412-F1-predicted_aligned_error_v6.png","plddt_mean":64.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=H1-4","jax_strain_url":"https://www.jax.org/strain/search?query=H1-4"},"sequence":{"accession":"P10412","fasta_url":"https://rest.uniprot.org/uniprotkb/P10412.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P10412/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P10412"}},"corpus_meta":[{"pmid":"16127177","id":"PMC_16127177","title":"HP1 binds specifically to Lys26-methylated histone H1.4, whereas simultaneous Ser27 phosphorylation blocks HP1 binding.","date":"2005","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16127177","citation_count":186,"is_preprint":false},{"pmid":"22465951","id":"PMC_22465951","title":"A dual role of linker histone H1.4 Lys 34 acetylation in transcriptional activation.","date":"2012","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/22465951","citation_count":75,"is_preprint":false},{"pmid":"21511733","id":"PMC_21511733","title":"Isoform-specific phosphorylation of human linker histone H1.4 in mitosis by the kinase Aurora B.","date":"2011","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/21511733","citation_count":54,"is_preprint":false},{"pmid":"23289424","id":"PMC_23289424","title":"Crosstalk between SET7/9-dependent methylation and ARTD1-mediated ADP-ribosylation of histone H1.4.","date":"2013","source":"Epigenetics & chromatin","url":"https://pubmed.ncbi.nlm.nih.gov/23289424","citation_count":50,"is_preprint":false},{"pmid":"31447100","id":"PMC_31447100","title":"Aberrant Function of the C-Terminal Tail of HIST1H1E Accelerates Cellular Senescence and Causes Premature Aging.","date":"2019","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/31447100","citation_count":48,"is_preprint":false},{"pmid":"29704315","id":"PMC_29704315","title":"Epigenetics and autism spectrum disorder: A report of an autism case with mutation in H1 linker histone HIST1H1E and literature review.","date":"2018","source":"American journal of medical genetics. 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mutagenesis of S27\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct in vitro binding with domain mapping and mutagenesis, replicated in later studies confirming the same mechanism\",\n      \"pmids\": [\"16127177\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Aurora B kinase phosphorylates H1.4 specifically at Ser27 in a cell-cycle-regulated manner peaking at metaphase; adjacent K26 dimethylation modulates Aurora B activity toward S27; H1.4 is the only somatic linker histone variant targeted by Aurora B; S27 phosphorylation increases H1.4 mobility and reduces chromatin binding in mitosis.\",\n      \"method\": \"In vitro kinase assays; in vivo phosphorylation analysis by immunofluorescence and immunoblot; FRAP on H1.4 mutants; cell-cycle fractionation\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — in vitro kinase assay plus FRAP in vivo, multiple orthogonal methods in single study\",\n      \"pmids\": [\"21511733\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Protein kinase A (PKA) phosphorylates H1.4 at Ser35; this phosphorylation causes H1.4 to dissociate from mitotic chromatin, and an H1.4-S35A mutant cannot rescue mitotic defects after H1.4 depletion; inhibition of PKA increases mitotic chromatin compaction in an H1.4-dependent manner.\",\n      \"method\": \"Mass spectrometry identification of phosphorylation site; in vitro kinase assay; mutant rescue experiments; PKA inhibitor treatment with chromatin compaction readout\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — in vitro kinase assay plus mutant rescue plus pharmacological inhibition, multiple orthogonal methods\",\n      \"pmids\": [\"21852232\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"GCN5 acetylates H1.4 at Lys34 (H1.4K34ac); this mark is enriched at promoters of active genes, increases H1.4 mobility (measured by FRAP), and recruits a general transcription factor to stimulate transcription.\",\n      \"method\": \"In vitro acetylation assay with GCN5; ChIP-seq for H1.4K34ac; FRAP of H1.4 mutants; transcription factor recruitment assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Moderate — in vitro enzymatic assay combined with ChIP, FRAP, and functional transcription data in a single study\",\n      \"pmids\": [\"22465951\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"SET7/9 methylates H1.4 at multiple lysine residues within KAK motifs in the C-terminal domain (K121, K129, K159, K171, K177, K192); ARTD1/PARP1-mediated poly(ADP-ribosyl)ation of H3 enables subsequent SET7/9-dependent methylation of H1.4; H1.4 and H3 compete for SET7/9 methylation.\",\n      \"method\": \"In vitro methylation assays with isolated histones; poly(ADP-ribosyl)ation assays; mass spectrometry identification of methylation sites\",\n      \"journal\": \"Epigenetics & chromatin\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro biochemical reconstitution but single lab, no in vivo validation reported\",\n      \"pmids\": [\"23289424\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Histone H1e (H1.4) physically interacts with the small hepatitis delta antigen (SHDAg) via H1.4's central globular domain; this interaction requires SHDAg oligomerization; N- or C-terminal deletion mutants of H1e inhibit HDV replication, which is rescued by wild-type H1e.\",\n      \"method\": \"Tandem affinity purification/mass spectrometry; domain-mapping by deletion mutagenesis; HDV replication rescue assays\",\n      \"journal\": \"Virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — TAP-MS identification plus domain mapping and functional rescue, single lab\",\n      \"pmids\": [\"18314153\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Frameshift mutations in the C-terminal tail of H1.4 produce stable nuclear proteins that bind chromatin, disrupt proper DNA compaction, are associated with a specific methylation pattern, reduce cell proliferation and S-phase entry, and accelerate cellular senescence.\",\n      \"method\": \"Expression of mutant H1.4 in cells; nuclear fractionation; DAPI staining for chromatin compaction; BrdU/flow cytometry for cell cycle; senescence assays; methylome analysis\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple cellular assays with defined molecular readouts, single lab\",\n      \"pmids\": [\"31447100\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Loss of H1.4 (and H1.2) in human PLB-985 cells induces an eosinophil-like transcriptional program and negatively regulates neutrophil lineage differentiation; this subtype-specific effect is confirmed in murine bone marrow stem cells.\",\n      \"method\": \"Genome-wide CRISPR/Cas9 screen; systematic disruption of individual H1 subtypes; transcriptional profiling; murine bone marrow differentiation assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with transcriptomic readout in two systems (human cell line and murine primary cells), single lab\",\n      \"pmids\": [\"32391789\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Phosphorylated H1.4 at serine 187 (pS187-H1.4) remains associated with active promoters genome-wide (enriched at transcription start sites of estrogen-activated genes), stably interacts with RNA Polymerase II, and is proposed to be phosphorylated by CDK9 as an early event in gene activation.\",\n      \"method\": \"ChIP-seq with pS187-H1.4-specific antibodies in MCF7 cells; co-immunoprecipitation with RNAPII; estradiol induction system\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genome-wide ChIP-seq with specific antibodies plus co-IP with RNAPII, single lab\",\n      \"pmids\": [\"33238524\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Exogenous expression of H1.4 C-terminal frameshift mutant (H1.4 CFT) in rat hippocampal neurons alters expression of ~400 genes (downregulating synaptic communication and neuropeptide signaling genes) and reduces neuronal activity as measured by multielectrode arrays.\",\n      \"method\": \"Exogenous expression of WT or mutant H1.4 in rat hippocampal neurons; genome-wide transcriptome analysis (RNA-seq); multielectrode array recording\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — transcriptome plus functional electrophysiology, single lab\",\n      \"pmids\": [\"34788807\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"A Rahman syndrome frameshift mutation in H1.4's C-terminal domain causes more extended/flexible nucleosome array conformation, enhanced linker DNA accessibility, inability to form compact stacked nucleosome structures, reduced capacity for liquid-liquid and liquid-solid phase separation, and increased H1.4 nuclear mobility; molecular dynamics simulations show the mutated CTD interacts with a shorter linker DNA segment.\",\n      \"method\": \"In vitro reconstitution of hexanucleosomal arrays; cryo-EM/structural analysis; FRET measurements; molecular dynamics simulations; FRAP in cells; micrococcal nuclease digestion\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — reconstitution with structural analysis, MD simulations, FRET, and FRAP, multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"42173878\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"H1.4 ablation causes robust changes in nascent transcription and gene expression; loss of H1.4 alters chromatin accessibility at enhancer-sized extragenic and intronic regions with concordant changes in H3K27ac/H3K4me1; sites losing accessibility are enriched for AP-1 motifs, placing H1.4 as a regulator of AP-1-directed enhancers.\",\n      \"method\": \"H1.4 knockout; nascent transcription assays (PRO-seq or equivalent); ATAC-seq; ChIP for H3K27ac and H3K4me1; motif analysis\",\n      \"journal\": \"iScience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO with ATAC-seq and histone modification ChIP, multiple orthogonal methods, single lab\",\n      \"pmids\": [\"41054521\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1994,\n      \"finding\": \"H1e (H1.4) binds preferentially and cooperatively to GC-rich DNA; a 25-residue C-terminal domain peptide retains GC-rich sequence preference but not cooperativity.\",\n      \"method\": \"DNA melting analysis; gel-mobility-shift assay; synthetic peptide binding experiments\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro biochemical assays with purified protein and synthetic peptides, single lab\",\n      \"pmids\": [\"8286360\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1996,\n      \"finding\": \"H1e (H1.4) is the only somatic linker histone variant that forms H1-H1 polymers when bound to oligonucleosomal DNA; poly(ADP-ribosyl)ation of H1e reduces polymer size without displacing it from linker regions.\",\n      \"method\": \"Cross-linking analysis; reverse-phase HPLC purification of H1 variants; methyl-accepting ability assay in native nuclei\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — biochemical reconstitution with purified variants and cross-linking, single lab\",\n      \"pmids\": [\"8687390\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"H1e (H1.4) variants specifically inhibit in vitro enzymatic DNA methylation and preferentially bind unmethylated CpG-rich DNA compared to H1c; the C-terminal domain mediates preferential binding to CpG-rich sequences.\",\n      \"method\": \"In vitro DNA methylation assay; gel retardation; Southwestern blot with CpG-rich oligonucleotides; competition binding assays\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro enzymatic and binding assays, single lab, replicated in companion 1996 paper\",\n      \"pmids\": [\"7848272\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"CYP1B1 physically associates with linker histone H1.4 (identified by LC-MS); downregulation of H1.4 is associated with increased chromatin accessibility and higher cell viability after PARP inhibitor treatment in resistant ovarian cancer cells.\",\n      \"method\": \"LC-MS proteomics for CYP1B1 interactors; ATAC-seq; H1.4 knockdown with cell viability readout; micrococcal nuclease digestion\",\n      \"journal\": \"Drug resistance updates\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — LC-MS identification of interaction without reciprocal validation; functional link is correlative, single lab\",\n      \"pmids\": [\"39395328\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Simultaneous knockdown of H1.2, H1.3, H1.4, and H1.5 in K562 cells reverses silencing of unintegrated HIV-1 DNA, resulting in increased viral expression; this effect is specific to HIV-1 and not observed for MLV unintegrated DNA.\",\n      \"method\": \"siRNA knockdown of four H1 variants; RT-qPCR/reporter assay for unintegrated HIV-1 and MLV expression\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — combined H1 subtype knockdown does not isolate H1.4-specific contribution; preprint, single method\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"SETDB1 binds to SUMOylated histone H1.2 and H1.4; SUMOylated H1.4 colocalizes with H3K9me3 at repetitive regions of the genome.\",\n      \"method\": \"PLAMseq (TurboID proximity labeling coupled to mass spectrometry and sequencing); SUMOylation mapping\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — proximity labeling approach without reciprocal validation; preprint, single method\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"The N-terminal domain peptide of histone H1e (residues 15-36) adopts two amphipathic alpha-helices separated by a flexible Gly-Gly motif under helix-stabilizing conditions; the Gly-Gly motif allows a wide range of relative orientations potentially facilitating DNA backbone tracking or simultaneous binding of two DNA segments.\",\n      \"method\": \"CD spectroscopy; 1H-NMR structure calculation in TFE solution\",\n      \"journal\": \"Protein science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — NMR structure determination with CD validation, but performed on isolated peptide in non-physiological conditions, single lab\",\n      \"pmids\": [\"11790831\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"Histone H1.4 is a linker histone that compacts chromatin by binding linker DNA (preferentially GC-rich sequences) and forming H1-H1 polymers on nucleosomal arrays; its functions are regulated by an array of post-translational modifications—including Aurora B-mediated S27 phosphorylation (which evicts HP1 bound to dimethyl-K26 and increases H1.4 mobility at mitosis), PKA-mediated S35 phosphorylation (which dissociates H1.4 from mitotic chromatin), CDK9/S187 phosphorylation (which keeps H1.4 at active promoters and promotes interaction with RNA Pol II), GCN5-mediated K34 acetylation (which increases mobility and recruits general transcription factors to active genes), and SET7/9-mediated C-terminal lysine methylation (modulated by ARTD1/PARP1 activity); disease-associated frameshift mutations in the C-terminal domain disrupt chromatin compaction, phase separation, and gene regulation, causing the neurodevelopmental Rahman syndrome.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"Histone H1.4 is a somatic linker histone that compacts chromatin by binding linker DNA—preferentially GC-rich and unmethylated CpG-rich sequences through its C-terminal domain—and is the only somatic H1 variant that forms H1-H1 polymers on oligonucleosomal arrays [#12, #13, #14]. Functional reconstitution shows that H1.4 drives formation of compact, stacked nucleosome structures and supports liquid-liquid/liquid-solid phase separation, with its C-terminal tail engaging linker DNA to limit accessibility [#10]. Its chromatin-binding behavior is tuned by a dense array of post-translational modifications: Aurora B phosphorylates Ser27 in a metaphase-peaking manner to increase H1.4 mobility and evict HP1, which otherwise reads dimethyl-Lys26 via a phospho-switch [#0, #1]; PKA phosphorylates Ser35 to dissociate H1.4 from mitotic chromatin [#2]; GCN5 acetylates Lys34 at active promoters to increase mobility and recruit general transcription factors [#3]; CDK9-dependent Ser187 phosphorylation keeps H1.4 at active transcription start sites in stable association with RNA Polymerase II [#8]; and SET7/9 methylates C-terminal KAK-motif lysines in a manner gated by PARP1/ARTD1 activity [#4]. Consistent with these roles, H1.4 loss reprograms nascent transcription and chromatin accessibility at AP-1-bound enhancers and biases hematopoietic differentiation [#11, #7]. C-terminal frameshift mutations that produce stable but functionally aberrant nuclear H1.4 cause the neurodevelopmental Rahman syndrome; these mutants fail to compact chromatin or phase separate, reduce proliferation and accelerate senescence, and in neurons downregulate synaptic-signaling genes and reduce neuronal activity [#6, #9, #10].\",\n  \"teleology\": [\n    {\n      \"year\": 1994,\n      \"claim\": \"Established the intrinsic DNA-binding preference of H1.4, showing it recognizes GC-rich linker DNA cooperatively through its C-terminal domain rather than binding DNA indiscriminately.\",\n      \"evidence\": \"DNA melting, gel-shift, and synthetic C-terminal peptide binding assays\",\n      \"pmids\": [\"8286360\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural basis for sequence preference\", \"Cooperativity determinant outside the tested 25-residue peptide not mapped\"]\n    },\n    {\n      \"year\": 1995,\n      \"claim\": \"Linked H1.4 DNA-binding specificity to epigenetic state by showing it preferentially binds unmethylated CpG-rich DNA and inhibits DNA methylation in vitro, distinguishing it functionally from H1c.\",\n      \"evidence\": \"In vitro methylation, gel retardation, and Southwestern assays with CpG-rich oligonucleotides\",\n      \"pmids\": [\"7848272\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"In vitro only; no cellular demonstration of methylation regulation\", \"Mechanism of methyltransferase inhibition unresolved\"]\n    },\n    {\n      \"year\": 1996,\n      \"claim\": \"Identified H1.4 as the unique somatic variant capable of self-polymerization on nucleosomal arrays, implicating it in higher-order chromatin organization, and showed poly(ADP-ribosyl)ation modulates polymer size.\",\n      \"evidence\": \"Cross-linking, HPLC variant purification, and methyl-accepting assays in native nuclei\",\n      \"pmids\": [\"8687390\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Functional consequence of polymer formation for chromatin compaction not directly tested in vivo\", \"Polymerization interface not mapped\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Provided structural rationale for linker DNA engagement by showing the H1e N-terminal peptide forms two amphipathic helices joined by a flexible Gly-Gly hinge that could allow DNA tracking or dual-segment binding.\",\n      \"evidence\": \"CD spectroscopy and 1H-NMR structure determination in TFE\",\n      \"pmids\": [\"11790831\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Isolated peptide in non-physiological solvent\", \"No structure on nucleosomal substrate\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined a phospho-methyl switch on the H1.4 N-terminal tail in which HP1 reads dimethyl-Lys26 and adjacent Ser27 phosphorylation blocks it, coupling H1.4 modification to heterochromatin reader recruitment.\",\n      \"evidence\": \"In vitro peptide-binding assays with the HP1 Chromo domain and S27 mutagenesis\",\n      \"pmids\": [\"16127177\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinase responsible for S27 not identified in this study\", \"In vivo dynamics of the switch not measured here\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified the mitotic kinases acting on the H1.4 tail: Aurora B phosphorylates S27 (peaking at metaphase, modulated by K26 methylation) and PKA phosphorylates S35, both reducing H1.4 chromatin binding, connecting the phospho-switch to mitotic chromatin dynamics.\",\n      \"evidence\": \"In vitro kinase assays, immunofluorescence/immunoblot, FRAP, mutant rescue, and PKA inhibition with compaction readouts\",\n      \"pmids\": [\"21511733\", \"21852232\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphatases reversing these marks not identified\", \"Interplay between S27 and S35 phosphorylation not resolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Connected H1.4 to active transcription by showing GCN5-mediated K34 acetylation marks active promoters, increases H1.4 mobility, and recruits a general transcription factor, recasting linker histone modification as activating rather than purely repressive.\",\n      \"evidence\": \"In vitro acetylation, ChIP-seq for H1.4K34ac, FRAP, and transcription factor recruitment assays\",\n      \"pmids\": [\"22465951\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Identity of the recruited GTF interaction surface not fully defined\", \"Whether K34ac and other marks co-occur on the same molecule unknown\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Mapped SET7/9 methylation of multiple C-terminal KAK-motif lysines on H1.4 and showed PARP1/ARTD1 ribosylation of H3 gates this methylation, linking H1.4 modification to a broader PARP-dependent regulatory network.\",\n      \"evidence\": \"In vitro methylation and poly(ADP-ribosyl)ation assays with mass spectrometry site mapping\",\n      \"pmids\": [\"23289424\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No in vivo validation of the methylation sites\", \"Functional consequence of C-terminal methylation untested\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Demonstrated a host-pathogen role in which H1.4's globular domain binds oligomerized small hepatitis delta antigen and supports HDV replication, the first non-chromatin functional partner identified.\",\n      \"evidence\": \"TAP-MS, deletion domain mapping, and HDV replication rescue assays\",\n      \"pmids\": [\"18314153\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which H1.4 promotes HDV replication unclear\", \"Single lab, no reciprocal in vivo validation\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Established H1.4 C-terminal frameshift mutations as causative of a disease state, showing the stable mutant proteins bind chromatin but disrupt compaction, alter methylation, reduce proliferation, and accelerate senescence.\",\n      \"evidence\": \"Expression of mutant H1.4 with fractionation, DAPI compaction, cell-cycle, senescence, and methylome assays\",\n      \"pmids\": [\"31447100\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanistic link from compaction defect to senescence not fully resolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed two distinct in vivo roles: pS187-H1.4 (CDK9-dependent) marks active promoters and binds RNA Pol II, while loss of H1.4 biases myeloid differentiation toward an eosinophil-like program, showing subtype-specific regulatory function.\",\n      \"evidence\": \"pS187 ChIP-seq and RNAPII co-IP in MCF7; genome-wide CRISPR screen and transcriptional/differentiation profiling in PLB-985 and murine bone marrow\",\n      \"pmids\": [\"33238524\", \"32391789\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct demonstration that CDK9 phosphorylates S187 not shown\", \"How H1.4 loss selects lineage outcome mechanistically unclear\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Connected the H1.4 C-terminal mutant to the neuronal phenotype of disease by showing it dysregulates ~400 genes, downregulating synaptic and neuropeptide signaling, and reduces neuronal electrical activity.\",\n      \"evidence\": \"Exogenous WT/mutant H1.4 expression in rat hippocampal neurons with RNA-seq and multielectrode array recording\",\n      \"pmids\": [\"34788807\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct chromatin mechanism at affected neuronal genes not mapped\", \"Exogenous overexpression may not model endogenous dosage\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Placed H1.4 as a regulator of AP-1-directed enhancers, showing its loss alters chromatin accessibility and enhancer histone marks at extragenic/intronic sites with concordant nascent transcription changes.\",\n      \"evidence\": \"H1.4 knockout with PRO-seq, ATAC-seq, H3K27ac/H3K4me1 ChIP, and motif analysis\",\n      \"pmids\": [\"41054521\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether H1.4 directly opposes AP-1 binding or acts indirectly unclear\", \"Single lab\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Provided the structural and biophysical mechanism of Rahman syndrome, showing a C-terminal frameshift mutant produces extended nucleosome arrays, increased linker DNA accessibility, failed compaction, impaired phase separation, and higher nuclear mobility.\",\n      \"evidence\": \"In vitro hexanucleosome reconstitution, cryo-EM/structural analysis, FRET, molecular dynamics, FRAP, and MNase digestion\",\n      \"pmids\": [\"42173878\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How phase-separation loss translates to specific gene dysregulation not directly linked\", \"Effect of disease mutant on PTM-mediated regulation untested\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unknown how the combinatorial array of H1.4 post-translational modifications is integrated on individual molecules to switch between compaction, phase separation, and active-promoter functions, and how disease mutations perturb this integrated code in specific cell types.\",\n      \"evidence\": \"No single study in the corpus reconstitutes the full modification network on defined chromatin\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No combinatorial PTM analysis on single molecules\", \"Phosphatases/demethylases for H1.4 marks unidentified\", \"Cell-type-specific rules linking H1.4 dosage to gene programs undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [12, 14, 10]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [10, 13]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [1, 6, 10]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [6, 8]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [3, 10, 11]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [3, 8, 11]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [1, 2]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"HP1\", \"AURKB\", \"PRKACA\", \"GCN5\", \"SETD7\", \"RNA Pol II\", \"PARP1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}