Affinage

DNAJC24

DnaJ homolog subfamily C member 24 · UniProt Q6P3W2

Length
149 aa
Mass
17.1 kDa
Annotated
2026-04-28
22 papers in source corpus 10 papers cited in narrative 10 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

DNAJC24 (DPH4) is a type III J-protein cochaperone essential for diphthamide biosynthesis on eukaryotic translation elongation factor 2 (eEF2). Its two-domain architecture comprises a J-domain that stimulates Hsp70 ATPase activity and a CSL-domain that coordinates iron in a rubredoxin-like tetrahedral geometry, enabling redox/electron carrier activity required for the radical SAM reaction catalyzed by the Dph1–Dph2 complex (PMID:22367199, PMID:15485916). Loss of DNAJC24 abolishes diphthamide modification of eEF2, conferring resistance to diphtheria toxin and Pseudomonas exotoxin A while sensitizing cells to TNF-mediated apoptosis; its promoter is subject to CpG methylation-based silencing that reversibly controls diphthamide levels and toxin sensitivity (PMID:26261303, PMID:22509046). Beyond diphthamide biosynthesis, DNAJC24 regulates podocyte adhesion, and in cancer cells it interacts with PCNA and promotes proliferation through PI3K/AKT signaling (PMID:31566424, PMID:38713100).

Mechanistic history

Synthesis pass · year-by-year structured walk · 8 steps
  1. 2004 High

    Identification of DPH4/DNAJC24 as a conserved gene required for diphthamide biosynthesis established that a DnaJ-like protein with a CSL zinc-finger domain participates in this post-translational modification pathway.

    Evidence Genetic identification and functional complementation of yeast dph4 mutants; sequence homology to human DNAJC24

    PMID:15485916

    Open questions at the time
    • Biochemical mechanism of DPH4 contribution unknown
    • No structural information
    • Relationship to other DPH factors unresolved
  2. 2008 High

    Mouse knockout demonstrated that Dph4 is essential for diphthamide modification in mammalian cells and revealed that DPH4 localizes to the cytoskeleton independently of the DPH1–DPH2–DPH3 complex, indicating a non-overlapping subcellular role.

    Evidence Dph4 knockout mouse embryonic cells; diphtheria toxin resistance assay; reporter-tagged protein localization imaging

    PMID:18765564

    Open questions at the time
    • Functional significance of cytoskeletal localization unclear
    • Whether DPH4 physically interacts with the DPH1-DPH2 complex not tested
  3. 2012 High

    Structural and spectroscopic characterization revealed DPH4 as a two-domain protein whose CSL-domain binds iron in tetrahedral coordination with rubredoxin-like properties, functioning as a redox/electron carrier, while its J-domain stimulates Hsp70 — establishing the molecular mechanism by which DPH4 supports diphthamide biosynthesis.

    Evidence NMR solution structure; UV-visible and EPR spectroscopy; in vitro iron-binding and redox activity assays

    PMID:22367199

    Open questions at the time
    • Direct electron transfer to Dph1-Dph2 Fe-S cluster not reconstituted for human DPH4
    • In vivo iron loading mechanism unknown
    • Relationship between Hsp70 stimulation and diphthamide biosynthesis unclear
  4. 2012 High

    Epigenetic silencing of DPH4 via CpG island hypermethylation was shown to be a physiologically relevant mechanism for abolishing diphthamide biosynthesis and conferring immunotoxin resistance, reversible by demethylating agents.

    Evidence CpG methylation analysis, RT-PCR, ADP-ribosylation assay, and 5-azacytidine rescue in an ALL cell line

    PMID:22509046

    Open questions at the time
    • Frequency of DPH4 methylation-based silencing in primary tumors unknown
    • Whether other DPH genes are co-silenced not addressed
  5. 2015 High

    Complete DPH4 knockout in human cells confirmed loss of diphthamide and toxin resistance while uncovering that diphthamide-deficient cells are pre-sensitized to TNF-induced apoptosis via NF-κB and death receptor pathways, linking diphthamide status to cell survival signaling.

    Evidence CRISPR knockout in MCF7 cells; ADP-ribosylation assay; toxin sensitivity and apoptosis assays

    PMID:26261303

    Open questions at the time
    • Mechanism connecting unmodified eEF2 to NF-κB/death receptor activation not defined
    • Whether TNF sensitization is eEF2-dependent or a DPH4-autonomous effect unclear
  6. 2019 Medium

    DPH4 knockdown in podocytes increased adhesion and impaired cell spreading, establishing a role for diphthamide pathway components in regulating integrin-mediated adhesion beyond canonical translational fidelity.

    Evidence Genome-scale RNAi screen and CRISPR-Cas9 knockout in human podocytes; fibronectin adhesion assay

    PMID:31566424

    Open questions at the time
    • Whether adhesion phenotype is mediated through eEF2 translation defects or a DPH4-autonomous function unclear
    • In vivo podocyte/kidney phenotype not examined
  7. 2022 Medium

    DNAJC24 was found to promote HCC cell proliferation and motility, with its transcription upregulated by HSF2 under stress, and its knockdown impairing ammonia metabolism and autophagy — suggesting functions beyond diphthamide biosynthesis.

    Evidence siRNA knockdown in HCC cell lines; proliferation, migration, ammonia metabolism, and autophagy assays; HSF2 knockdown

    PMID:35606363

    Open questions at the time
    • Ammonia metabolism link is correlative; direct enzymatic target unknown
    • Whether proliferation effect is diphthamide-dependent not tested
    • Single lab, no independent replication
  8. 2024 Medium

    Identification of a direct DNAJC24–PCNA interaction and downstream PI3K/AKT activation in lung adenocarcinoma cells expanded the functional repertoire of DNAJC24 to include a signaling axis promoting tumor cell proliferation and invasion.

    Evidence Co-immunoprecipitation and mass spectrometry in A549 and NCI-H1299 cells; AKT phosphorylation assay; proliferation and invasion assays

    PMID:38713100

    Open questions at the time
    • PCNA interaction not validated by reciprocal Co-IP or structural methods
    • Whether PCNA interaction is J-domain or CSL-domain mediated unknown
    • Relationship between PCNA binding and diphthamide biosynthesis not addressed

Open questions

Synthesis pass · forward-looking unresolved questions
  • Key unresolved questions include: (1) whether DPH4 directly donates electrons to the Dph1-Dph2 Fe-S cluster in vivo, (2) the functional significance of its cytoskeletal localization, (3) whether its cancer-associated proliferative roles are diphthamide-dependent or reflect independent functions of its J-domain or PCNA interaction, and (4) whether Hsp70 cochaperone activity is required for diphthamide biosynthesis or other cellular processes.
  • No in vivo reconstitution of electron transfer from DPH4 to Dph1-Dph2
  • No crystal or cryo-EM structure of DPH4 in complex with partners
  • In vivo relevance of PCNA interaction and PI3K/AKT activation not established

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016491 oxidoreductase activity 2 GO:0044183 protein folding chaperone 2 GO:0098772 molecular function regulator activity 1
Localization
GO:0005829 cytosol 2 GO:0005856 cytoskeleton 1
Pathway
R-HSA-392499 Metabolism of proteins 4 R-HSA-162582 Signal Transduction 1
Partners
DPH1DPH2HSF2PCNA

Evidence

Reading pass · 10 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2004 DPH4 (DNAJC24) encodes a CSL zinc finger-containing DnaJ-like protein required for diphthamide biosynthesis on translation elongation factor 2 (EF2); human DPH4 has sequence homology to yeast Dph4 and functions in this conserved pathway. Genetic identification and functional complementation in yeast; sequence homology analysis Molecular and cellular biology High 15485916
2008 Mouse Dph4 (ortholog of human DNAJC24) is required for diphthamide modification of eEF2; cells from homozygous mutant embryos lacked diphthamide on eEF2 and were resistant to diphtheria toxin killing. Reporter-tagged DPH4 localized to the cytoskeleton, distinct from DPH1 localization, consistent with DPH4 not being part of the DPH1-DPH2-DPH3 complex. Mouse knockout model; diphtheria toxin resistance assay; reporter-tagged protein localization imaging Journal of cell science High 18765564
2012 Human DPH4 (DNAJC24) is a type III J-protein with an NMR-determined two-domain structure: a conserved J-domain and a CSL-domain connected by a flexible linker-helix. The CSL-domain binds iron in tetrahedral coordination via cysteines, forming an Fe-Dph4 species with rubredoxin-like spectral properties. Fe-Dph4 undergoes oligomerization and exhibits redox/electron carrier activity critical for diphthamide biosynthesis. Iron binding also enhances Hsp70-stimulatory (J-domain) function. NMR solution structure; UV-visible spectroscopy; EPR spectroscopy; in vitro iron-binding assay; redox activity assay The Journal of biological chemistry High 22367199
2014 Yeast Dph3 (ortholog of human DPH3/KTI11, a CSL zinc finger protein closely related to DPH4/DNAJC24) functions as an electron donor to reduce the Fe-S cluster in the Dph1-Dph2 complex during the first step of diphthamide biosynthesis, establishing that Dph3/Dph4-family CSL proteins serve as electron carriers in this pathway. In vitro reconstitution of diphthamide biosynthesis; EPR spectroscopy; iron-binding assay Journal of the American Chemical Society Medium 24422557
2015 Complete knockout of DPH4 in MCF7 cells generates viable cells with unmodified eEF2 (no diphthamide), rendering cells resistant to Pseudomonas exotoxin A and diphtheria toxin ADP-ribosylation, but pre-sensitizing cells to TNF-mediated apoptosis via NF-κB and death receptor pathway activation. CRISPR/gene knockout; ADP-ribosylation assay; toxin sensitivity assay; apoptosis assay Proceedings of the National Academy of Sciences of the United States of America High 26261303
2012 Immunotoxin resistance in an ALL cell line is caused by epigenetic silencing of DPH4 (DNAJC24) via CpG island hypermethylation of its promoter, reducing DPH4 mRNA and protein levels, which prevents diphthamide biosynthesis and renders EF2 refractory to ADP-ribosylation by Pseudomonas exotoxin A. CpG methylation analysis; RT-PCR; protein expression analysis; ADP-ribosylation assay; methylation inhibitor rescue (5-azacytidine) Proceedings of the National Academy of Sciences of the United States of America High 22509046
2019 Knockdown of DPH4 (DNAJC24) in human podocytes (via shRNA or CRISPR-Cas9 KO) increases adhesion to fibronectin and sFLT1/Fc substrates and causes a cell-spreading defect, establishing a role for diphthamide biosynthesis genes including DPH4 in regulating podocyte adhesion. Genome-scale RNAi screen; CRISPR-Cas9 knockout; adhesion assay; cell-spreading assay American journal of physiology. Renal physiology Medium 31566424
2021 CRISPR/Cas9 pooled screen in pig PK-15 cells confirmed DNAJC24 as a host factor essential for diphtheria toxin-induced cell death, with individual gene knockout validating that loss of DNAJC24 confers DT resistance by blocking diphthamide biosynthesis. Genome-scale CRISPR/Cas9 screen; individual gene knockout; diphtheria toxin cytotoxicity assay Biotechnology journal Medium 34705337
2022 DNAJC24 knockdown in HCC cells reduces proliferation and motility; its expression is transcriptionally upregulated by HSF2 under hypoxia, starvation, and heat stress. Targeting DNAJC24 under normal conditions affects HCC cell proliferation and autophagy by interfering with ammonia metabolism. siRNA knockdown; cell proliferation and migration assays; transcription factor knockdown (HSF2); ammonia metabolism assay; autophagy assay Cell death & disease Medium 35606363
2024 DNAJC24 directly interacts with PCNA (confirmed by Co-IP and mass spectrometry) and promotes lung adenocarcinoma (LUAD) cell proliferation and invasion by activating AKT phosphorylation via the PI3K/AKT signaling pathway. Co-immunoprecipitation; mass spectrometry; AKT phosphorylation assay; cell proliferation and invasion assays in A549 and NCI-H1299 cell lines FASEB journal Medium 38713100

Source papers

Stage 0 corpus · 22 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2004 Identification of the proteins required for biosynthesis of diphthamide, the target of bacterial ADP-ribosylating toxins on translation elongation factor 2. Molecular and cellular biology 145 15485916
2007 Network of general and specialty J protein chaperones of the yeast cytosol. Proceedings of the National Academy of Sciences of the United States of America 126 17438278
2014 Dph3 is an electron donor for Dph1-Dph2 in the first step of eukaryotic diphthamide biosynthesis. Journal of the American Chemical Society 58 24422557
2008 Diphthamide modification of eEF2 requires a J-domain protein and is essential for normal development. Journal of cell science 57 18765564
2015 Loss of diphthamide pre-activates NF-κB and death receptor pathways and renders MCF7 cells hypersensitive to tumor necrosis factor. Proceedings of the National Academy of Sciences of the United States of America 49 26261303
2012 Immunotoxin resistance via reversible methylation of the DPH4 promoter is a unique survival strategy. Proceedings of the National Academy of Sciences of the United States of America 43 22509046
2006 A genomewide analysis of genes for the heat shock protein 70 chaperone system in the ascidian Ciona intestinalis. Cell stress & chaperones 30 16572726
2012 Structure and mechanistic insights into novel iron-mediated moonlighting functions of human J-protein cochaperone, Dph4. The Journal of biological chemistry 27 22367199
2012 Synthetic lethal interactions in yeast reveal functional roles of J protein co-chaperones. Molecular bioSystems 26 22851130
2011 A 556 kb deletion in the downstream region of the PAX6 gene causes familial aniridia and other eye anomalies in a Chinese family. Molecular vision 25 21321669
2015 Protective effect of the multitarget compound DPH-4 on human SSAO/VAP-1-expressing hCMEC/D3 cells under oxygen-glucose deprivation conditions: an in vitro experimental model of cerebral ischaemia. British journal of pharmacology 14 26362823
2021 Establishment of a pig CRISPR/Cas9 knockout library for functional gene screening in pig cells. Biotechnology journal 13 34705337
2022 DNAJC24 is a potential therapeutic target in hepatocellular carcinoma through affecting ammonia metabolism. Cell death & disease 12 35606363
2022 Genetic Analysis of HSP40/DNAJ Family Genes in Parkinson's Disease: a Large Case-Control Study. Molecular neurobiology 11 35715682
2023 Screening of reliable reference genes for the normalization of RT-qPCR in chicken gastrointestinal tract. Poultry science 10 37918133
2019 Forward genetic screen in human podocytes identifies diphthamide biosynthesis genes as regulators of adhesion. American journal of physiology. Renal physiology 8 31566424
2015 A familial pericentric inversion of chromosome 11 associated with a microdeletion of 163 kb and microduplication of 288 kb at 11p13 and 11q22.3 without aniridia or eye anomalies. American journal of medical genetics. Part A 8 26419218
2018 Deletion distal to the PAX6 coding region reveals a novel basis for familial cosegregation of aniridia and diabetes mellitus. Diabetes research and clinical practice 7 30572005
2025 A genome-wide association study identified candidate genes associated with egg quality traits in Muscovy duck. BMC genomics 5 40301754
2024 DNAJC24 acts directly with PCNA and promotes malignant progression of LUAD by activating phosphorylation of AKT. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 3 38713100
2023 Elucidation of novel SNPs affecting immune response to classical swine fever vaccination in pigs using immunogenomics approach. Veterinary research communications 3 38017322
2024 Arabidopsis Dph4 is an Hsp70 Cochaperone with Iron-Binding Properties. ACS omega 2 39281955