Affinage

CHCHD4

Mitochondrial intermembrane space import and assembly protein 40 · UniProt Q8N4Q1

Length
142 aa
Mass
16.0 kDa
Annotated
2026-04-28
80 papers in source corpus 42 papers cited in narrative 40 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

CHCHD4 (human MIA40) is the central oxidoreductase and import receptor of the mitochondrial intermembrane space (IMS) disulfide relay system, coupling protein translocation, oxidative folding, and chaperone-mediated stabilization of nuclear-encoded IMS precursors. Its folded core comprises an α-helical hairpin stabilized by two structural twin-CX9C disulfides, with a CPC active-site motif adjacent to a hydrophobic substrate-binding cleft that captures incoming precursors bearing a 9-residue IMS-targeting signal, forms transient mixed disulfides to introduce native disulfide bonds, and can also act as a disulfide isomerase that proofreads non-native linkages (PMID:19182799, PMID:25451030, PMID:20026652). CHCHD4 is reoxidized by ALR/Erv1, which shuttles electrons to cytochrome c, forming a thermodynamically ordered relay (substrate → CHCHD4 → ALR → cytochrome c), and ALR itself controls CHCHD4 mitochondrial accumulation (PMID:19477928, PMID:23186364). Beyond oxidoreductase activity, CHCHD4 functions as a holdase chaperone that stabilizes cysteine-free substrates (e.g., HAX1, Atp23) through its hydrophobic cleft, physically interacts with AIFM1—which promotes CHCHD4 import and whose loss reduces respiratory chain complex assembly—and regulates HIF-1α stabilization, intracellular oxygenation, and perinuclear mitochondrial positioning through its control of respiratory chain activity (PMID:22990235, PMID:39564806, PMID:26004228, PMID:22214851, PMID:28497026).

Mechanistic history

Synthesis pass · year-by-year structured walk · 16 steps
  1. 2004 High

    Discovery that Mia40 is an essential mitochondrial IMS import factor for small cysteine-rich proteins resolved how these substrates, which lack classical presequences, are recognized and retained in the IMS.

    Evidence Yeast mia40 mutant with in organello import assays and co-immunoprecipitation showing selective import defect for small Tim proteins

    PMID:15359280

    Open questions at the time
    • Human ortholog function not yet tested
    • Mechanism of substrate recognition undefined
    • Relationship to oxidative folding unknown
  2. 2005 High

    Establishing the human disulfide relay: CHCHD4 was shown to localize to the IMS as a disulfide-bonded protein whose depletion selectively reduces IMS substrates, while Erv1 was identified as its dedicated reoxidant linking the pathway to the respiratory chain.

    Evidence siRNA knockdown of human MIA40, redox trapping, and parallel yeast Erv1 temperature-sensitive mutant studies with cytochrome c electron transfer assays

    PMID:16181637 PMID:16185707 PMID:16185709

    Open questions at the time
    • Structural basis of substrate binding unknown
    • Electron flow thermodynamics not quantified
  3. 2007 High

    Identification of a single N-terminal substrate cysteine as the docking residue for mixed-disulfide formation with Mia40 defined the site-specific trans-receptor mechanism and separated translocation from downstream assembly steps.

    Evidence Systematic cysteine mutagenesis of Tim9/Tim10 with in organello and in vitro import assays and disulfide trapping

    PMID:17553782 PMID:17680986

    Open questions at the time
    • Structural basis of specificity not yet resolved
    • Role of hydrophobic interactions in recognition not tested
  4. 2008 High

    Mutagenesis of the CPC motif dissected the dual function of its cysteines: one mediates catalytic disulfide exchange with both substrate and Erv1, while the twin-CX9C cysteines are purely structural, clarifying which residues are enzymatically active.

    Evidence Cysteine mutagenesis with yeast viability assays and in vitro reconstitution of Mia40–Erv1–Tim10 pathway

    PMID:19011240

    Open questions at the time
    • Atomic structure not yet available
    • Substrate-binding cleft contribution not defined
  5. 2009 High

    Atomic structures of human MIA40 (NMR) and yeast Mia40 (crystal), combined with full in vitro reconstitution of the oxidative folding pathway with measured redox midpoint potentials, established the complete structural and thermodynamic framework for the disulfide relay: a hydrophobic cleft captures a 9-residue IMS-targeting signal helix, positioning the substrate cysteine for mixed-disulfide formation, with electron flow driven by redox potential differences.

    Evidence NMR (human) and X-ray crystallography (yeast) of the core domain; hydrophobic cleft mutagenesis with yeast complementation; in vitro reconstitution with Tim13 and midpoint potential measurements; ITS mutagenesis and import assays

    PMID:19182799 PMID:19477928 PMID:19667201 PMID:20026652

    Open questions at the time
    • Chaperone function beyond oxidative trapping not yet recognized
    • In vivo redox state regulation unknown
  6. 2010 High

    NMR characterization of folding intermediates revealed that Mia40 acts as a chaperone that induces α-helical folding of the substrate ITS before disulfide bond formation, establishing a two-step induced-folding mechanism for oxidative trapping.

    Evidence NMR structural characterization of substrate folding intermediates in complex with Mia40

    PMID:21059946

    Open questions at the time
    • Whether chaperone activity extends to non-cysteine substrates unknown
    • Folding kinetics not quantified
  7. 2011 High

    Crystal structure of the covalent ALR–MIA40 mixed-disulfide intermediate defined how electrons transfer from MIA40 to ALR, with ALR's N-terminal domain mimicking substrate binding to the hydrophobic cleft, and expanded the substrate repertoire to include non-twin-CX9C proteins like anamorsin.

    Evidence X-ray crystallography of ALR–MIA40 covalent complex; in vitro disulfide bond formation and NMR characterization with anamorsin as substrate

    PMID:21383138 PMID:21700214

    Open questions at the time
    • Full range of human substrates not cataloged
    • Whether ALR and substrate compete for the same binding site in vivo unclear
  8. 2012 High

    Three parallel advances broadened the Mia40 paradigm: discovery of a disulfide-independent chaperone activity for Atp23 folding, identification of ternary Erv1–Mia40–substrate complexes ensuring full substrate oxidation, and linking CHCHD4 to HIF-1α stabilization and tumor growth through modulation of oxygen consumption.

    Evidence Cysteine-free Atp23 import reconstitution; in vivo ternary complex disulfide trapping; siRNA/overexpression of CHCHD4 with tumor xenografts and oxygen consumption measurements

    PMID:22214851 PMID:22918950 PMID:22990235

    Open questions at the time
    • Direct mechanism linking CHCHD4 to HIF-1α not defined at molecular level
    • Range of chaperone-only substrates unknown
  9. 2014 High

    Kinetic reconstitution with Cox17 demonstrated that Mia40 is not just an oxidase but also a disulfide isomerase that proofreads non-native disulfides, with the hydrophobic cleft selecting the reactive substrate cysteine and controlling mixed-disulfide lifetime.

    Evidence Stopped-flow kinetic analysis and disulfide mapping of Cox17 oxidative folding intermediates with Mia40

    PMID:24407114 PMID:25451030

    Open questions at the time
    • Isomerase activity not tested for substrates beyond Cox17
    • In vivo relevance of proofreading not demonstrated
  10. 2015 High

    The AIFM1–CHCHD4 functional axis was established: AIFM1 promotes CHCHD4 mitochondrial import, CHCHD4 depletion recapitulates AIFM1-deficiency respiratory defects, and forced CHCHD4 mitochondrial targeting rescues respiratory chain assembly in AIFM1-deficient cells, while Mia40's role extended to catalyzing the MICU1–MICU2 disulfide required for mitochondrial Ca²⁺ uptake regulation.

    Evidence Reciprocal Co-IP, siRNA, rescue by forced mitochondrial CHCHD4 localization, embryoid body assays; MICU1/MICU2 in vitro disulfide formation plus Ca²⁺ uptake measurements

    PMID:26004228 PMID:26158520 PMID:26387864

    Open questions at the time
    • Structural basis of AIFM1–CHCHD4 interaction not yet resolved
    • How AIFM1 mechanistically promotes CHCHD4 import unclear
  11. 2016 High

    Domain dissection established that the hydrophobic substrate-binding cleft is both necessary and sufficient for import (acting as a 'holding trap' trans-site receptor), while the CPC oxidase domain, though required for viability, could be partially replaced by chemical oxidant, separating receptor and redox functions.

    Evidence Domain-deletion mutagenesis with yeast complementation and diamide chemical rescue

    PMID:27343349

    Open questions at the time
    • Whether receptor and oxidase functions are temporally separable in vivo unknown
  12. 2017 Medium

    CHCHD4 was shown to control perinuclear mitochondrial positioning in hypoxia through its CPC-dependent respiratory chain activity, linking the disulfide relay to HIF-1α activation and providing a mechanism for how CHCHD4 influences oxygen sensing.

    Evidence Live mitochondrial imaging with CPC mutagenesis, CHCHD4 overexpression/knockdown, complex IV inhibition, and HIF-1α siRNA

    PMID:28497026

    Open questions at the time
    • Direct molecular link between respiratory chain activity and mitochondrial positioning not identified
    • Whether mitochondrial positioning is causal or correlative for HIF activation not settled
  13. 2020 Medium

    S-glutathionylation of MIA40's structural cysteines was identified as a regulatory post-translational modification affecting complex III/IV activities and enabling direct electron transfer to cytochrome c, revealing a redox-regulatory layer beyond simple oxidized/reduced cycling.

    Evidence MALDI MS of in vivo protein, glutathionylation-deficient mutagenesis, immunocapture of complex III, electron transfer assays

    PMID:32971361

    Open questions at the time
    • Physiological triggers of glutathionylation not identified
    • Fe-S cluster involvement in electron transfer not independently confirmed
    • Stoichiometry and dynamics of glutathionylation in vivo unknown
  14. 2023 Medium

    A CHCHD4–TRIAP1–cardiolipin axis was defined linking CHCHD4 to innate immune signaling: exercise-induced CHCHD4 downregulation reduces TRIAP1 import, decreasing cardiolipin, promoting VDAC oligomerization and mtDNA release, and activating cGAS-STING/NF-κB to drive slow-twitch muscle fiber specification.

    Evidence CHCHD4 haploinsufficient mice, TRIAP1 import assays, cardiolipin measurement, cGAS-STING pathway activation assays

    PMID:38157298

    Open questions at the time
    • Whether TRIAP1 is the sole mediator of the cardiolipin effect not tested
    • Pathway not reconstituted outside skeletal muscle context
  15. 2024 High

    Crystal structure of AIF bound to the CHCHD4 N-terminal peptide resolved the structural basis of the AIFM1–CHCHD4 interaction at the AIF active site, revealing allosteric effects on cofactor binding and explaining how AIFM1 promotes CHCHD4 import.

    Evidence X-ray crystallography of AIF–NAD⁺–FAD–CHCHD4 peptide complex, cross-linking mass spectrometry, site-directed mutagenesis

    PMID:38460521

    Open questions at the time
    • Structure captures only N-terminal peptide, not full-length CHCHD4
    • How allosteric changes translate to enhanced import not mechanistically resolved
  16. 2025 High

    Quantitative oxidative folding reconstitution showed MIA40 accelerates TRIAP1 folding 30-fold by bypassing a kinetic disulfide trap, and the AIFM1–MIA40 complex was shown to suppress AIFM1-mediated cell death in a NADH-dependent manner, linking metabolic state to apoptotic threshold.

    Evidence In vitro oxidative folding kinetics with NMR and MS for TRIAP1; complex I KO cells with MIA40 siRNA, NADH depletion, and cell death assays for AIFM1 regulation

    PMID:39909379 PMID:40055465

    Open questions at the time
    • Whether NADH-dependent complex regulation is universal across cell types unknown
    • Full scope of MIA40's anti-apoptotic role via AIFM1 sequestration not defined

Open questions

Synthesis pass · forward-looking unresolved questions
  • Key unresolved questions include: the complete human substrate repertoire of CHCHD4, the molecular mechanism by which CHCHD4/respiratory chain activity drives perinuclear mitochondrial positioning, the physiological significance of the Fe-S cluster-bound form, and how CHCHD4 redox state is dynamically regulated in response to cellular stress.
  • Comprehensive substrate catalogue lacking
  • Fe-S cluster function undefined
  • Mitochondrial positioning mechanism not molecularly resolved
  • In vivo redox regulation poorly understood

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0140096 catalytic activity, acting on a protein 9 GO:0016491 oxidoreductase activity 4 GO:0044183 protein folding chaperone 3 GO:0016853 isomerase activity 1
Localization
GO:0005739 mitochondrion 5
Pathway
R-HSA-392499 Metabolism of proteins 9 R-HSA-9609507 Protein localization 6 R-HSA-5357801 Programmed Cell Death 1
Partners
AIFM1CHCHD10CYCSGFERHAX1MICU1MICU2TRIAP1
Complex memberships
AIFM1-CHCHD4 complexMIA40-ALR/Erv1 disulfide relay

Evidence

Reading pass · 40 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2004 Mia40 (yeast ortholog of CHCHD4) is an essential component of the mitochondrial IMS protein import machinery; mitochondria with mutant Mia40 are selectively impaired in import of small IMS proteins (Tim9, Tim10), and Mia40 directly binds small Tim proteins as an initial step in their assembly into IMS complexes. Yeast genetics (mia40 mutant), in organello import assays, co-immunoprecipitation/binding assays The EMBO journal High 15359280
2005 Human MIA40 (CHCHD4) localizes as a soluble protein in the mitochondrial IMS where it forms complexes; depletion by RNAi specifically reduces steady-state levels of small cysteine-containing IMS proteins (DDP1, TIM10A); import and stability of MIA40 itself depends on conserved twin CX9C cysteine residues that form intramolecular disulfide bonds. siRNA knockdown, subcellular fractionation, thiol-trapping redox assays, cysteine mutagenesis Journal of molecular biology High 16185709
2005 Erv1 (yeast sulfhydryl oxidase/ALR) cooperates with Mia40 in the biogenesis of small IMS proteins; erv1-ts mitochondria show selective import/assembly defects for small Tims; Erv1 associates with Mia40 in a reductant-sensitive (disulfide-bonded) manner and functionally links the Mia40 import pathway to the respiratory chain by shuttling electrons to cytochrome c. Temperature-sensitive yeast mutants, in organello import assays, thiol trapping, biochemical interaction assays Journal of molecular biology High 16181637 16185707
2007 Mia40 acts as a site-specific trans-receptor for IMS substrates: only the most N-terminal cysteine of Tim9/Tim10 precursors is critical for translocation across the outer membrane and mixed-disulfide formation with Mia40; subsequent cysteines drive release and assembly. Systematic cysteine mutagenesis, in organello and in vitro import assays, disulfide trapping The Journal of biological chemistry High 17553782 17680986
2008 The CPC motif of Mia40 contains functionally distinct cysteines: the second cysteine (C55 in human) is essential for viability and forms a mixed disulfide with Erv1; the twin CX9C cysteines are structural, stabilizing the folded domain; the CPC disulfide mediates redox reactions with both Erv1 and substrate proteins in a reconstituted system. Cysteine mutagenesis, yeast growth assays, in vitro reconstitution with Mia40 + Erv1 + Tim10, disulfide trapping The Journal of biological chemistry High 19011240
2009 Solution NMR structure of human MIA40 (CHCHD4) reveals a 66-residue folded domain with an alpha-helical hairpin core stabilized by two structural disulfides and a CPC active site adjacent to a hydrophobic substrate-binding cleft; the second CPC cysteine (Cys55) is essential for mixed disulfide formation with substrate; mutations in the hydrophobic cleft are lethal in vivo and abolish substrate binding in vitro. NMR structure determination, active-site mutagenesis, in vitro binding assays, yeast complementation Nature structural & molecular biology High 19182799
2009 Crystal structure of yeast Mia40 core domain reveals a fruit-dish shape with a hydrophobic concave region that accommodates substrate in a helical conformation; the CPC disulfide is adjacent to this binding site; mutation of hydrophobic residues causes growth defects and impaired substrate assembly. X-ray crystallography (3 Å), hydrophobic-residue mutagenesis, yeast growth and import assays Proceedings of the National Academy of Sciences of the United States of America High 19667201
2009 Mia40 substrates contain a 9-amino acid IMS-targeting signal (ITS) that forms an amphipathic helix; the ITS is sufficient for outer membrane crossing and docks onto the hydrophobic substrate-binding cleft of Mia40 via hydrophobic interactions (µM affinity), orienting the docking cysteine for mixed-disulfide formation in a two-step mechanism. ITS mutagenesis, in vitro binding assays, targeting/import assays with ITS-fusion proteins The Journal of cell biology High 20026652
2009 The Mia40-Erv1 oxidative folding pathway for small Tim proteins was fully reconstituted in vitro with Tim13 as substrate: Mia40 directly oxidizes Tim13 inserting two disulfide bonds sequentially; Erv1 reoxidizes Mia40; midpoint potentials establish electron flow Tim13 (−310 mV) → Mia40 (−290 mV) → Erv1 C130-C133 pair (−150 mV); the CPC cysteines of Mia40 are required for substrate oxidation. In vitro reconstitution, redox midpoint potential measurements, disulfide trapping, cysteine mutagenesis of Mia40 and Erv1 Molecular biology of the cell High 19477928
2010 Mia40 functions as a molecular chaperone assisting α-helical folding of the ITS of incoming substrates (first induced-folding step); the folded ITS then acts as a scaffold to drive folding of the second substrate helix in a Mia40-independent manner (second induced-folding step), constituting the oxidative protein-trapping mechanism. NMR structural characterization of substrate folding intermediates at each stage and in complex with Mia40 Proceedings of the National Academy of Sciences of the United States of America High 21059946
2011 ALR (human Erv1 ortholog) interacts with MIA40 via its unstructured N-terminal domain, which mimics substrate binding to the MIA40 hydrophobic cleft; crystal structure of the ALR-MIA40 covalent mixed-disulfide intermediate was determined, revealing the molecular basis of electron transfer from MIA40 to ALR. X-ray crystallography of covalent ALR-MIA40 complex, biochemical interaction assays, domain dissection Proceedings of the National Academy of Sciences of the United States of America High 21383138
2011 Human anamorsin (a [2Fe-2S] cluster protein) is a substrate of Mia40 (CHCHD4): Mia40 introduces two disulfide bonds in the twin CX2C motif of anamorsin's C-terminal domain via an intermolecular mixed-disulfide intermediate, enabling anamorsin import into the IMS. In vitro disulfide bond formation assays, NMR/structural characterization, disulfide trapping, mitochondrial import assays Chemistry & biology High 21700214
2011 Mia40 oxidizes cysteines 27 and 64 in domain I of Ccs1 (copper chaperone for Sod1), forming a structural disulfide that drives Ccs1 import into the IMS; these cysteines are dispensable for cytosolic Ccs1 function but essential for mitochondrial accumulation, establishing a Mia40-mediated mechanism that controls the cellular distribution of Ccs1 and consequently active Sod1. Cysteine mutagenesis, in organello import assays, thiol-trapping, co-immunoprecipitation Molecular biology of the cell High 21865594
2012 Atp23 is a novel substrate of Mia40 with a complex disulfide pattern (10 cysteines); Mia40 can fold Atp23 via its hydrophobic substrate-binding pocket independent of disulfide bond formation (a cysteine-free Atp23 variant still imports in a Mia40-dependent manner), revealing a chaperone-like folding activity of Mia40 beyond oxidative trapping. In vitro folding/aggregation assays, cysteine-to-serine mutagenesis, in organello import assays The EMBO journal High 22990235
2012 In vivo and in organello experiments show that Erv1 directly participates in Mia40-substrate complex dynamics by forming a ternary Erv1-Mia40-substrate complex, ensuring that substrate proteins are released from Mia40 in a fully oxidized (two-disulfide) form. In vivo disulfide trapping, in organello import assays, ternary complex detection Molecular biology of the cell High 22918950
2012 Human ALR (Erv1 ortholog) controls the mitochondrial localization of human MIA40 (CHCHD4) by mediating disulfide bond formation that enables its import; a disease-associated mutation in ALR impairs MIA40 accumulation in mitochondria. Complementation assays, mitochondrial import assays, redox state analysis Traffic (Copenhagen, Denmark) High 23186364
2012 CHCHD4 (MIA40) modulates cellular oxygen consumption rate and metabolism; elevated CHCHD4 expression promotes HIF-1α protein stabilization in hypoxia; CHCHD4 knockdown blocks HIF-1α induction and inhibits tumor growth and angiogenesis in vivo; the effect on HIF-1α is insensitive to antioxidant treatment. siRNA knockdown, stable overexpression, in vivo tumor xenograft assays, oxygen consumption measurements The Journal of clinical investigation High 22214851
2013 Mia40 is identified as an iron-sulfur protein: yeast Mia40 binds a [2Fe-2S] cluster as a dimer, coordinated by the CPC cysteine residues; cellular iron uptake analyses confirm in vivo iron binding; Fe-S cluster-containing Mia40 does not donate electrons to Erv1, suggesting a function distinct from its thiol oxidoreductase activity. In vitro and in vivo [2Fe-2S] cluster binding assays, EPR spectroscopy, iron uptake analysis, oxygen consumption measurement The Biochemical journal Medium 23834247
2013 Mia40 facilitates the biogenesis and complex assembly of Tim22, a multispanning inner membrane protein, by forming a disulfide-bonded intermediate with Tim22 and also via non-covalent interactions, extending Mia40's role beyond IMS-soluble proteins to inner membrane protein integration. In organello import assays, disulfide trapping, co-immunoprecipitation Molecular biology of the cell High 23283984
2014 Mia40 initially forms a dynamic non-covalent enzyme-substrate complex with Cox17 (µs–ms lifetime), then rapidly forms a mixed disulfide specifically with Cys36 of Cox17 because of neighboring hydrophobic residues; hydrophobic substrate binding drives selection of the reactive cysteine and the long lifetime of the mixed disulfide retains partially folded proteins for complete oxidation. Kinetic analysis of mixed disulfide formation, Cox17 mutagenesis, stopped-flow/rapid mixing assays Nature communications High 24407114
2014 Mia40 combines thiol oxidase and disulfide isomerase activity: it preferentially forms native disulfides in Cox17, acts as a proofreading disulfide reductase/isomerase to reshuffle non-native disulfides, driven by conformational folding of the substrate and the hydrophobic binding site of Mia40. In vitro oxidative folding assays with Cox17, kinetic analysis, disulfide bond mapping Journal of molecular biology High 25451030
2015 CHCHD4 (MIA40) physically interacts with AIF (AIFM1); AIF deficiency reduces CHCHD4 protein levels by diminishing its mitochondrial import (post-transcriptionally); CHCHD4 depletion alone recapitulates the respiratory chain defect of AIF-deficient cells; restoring CHCHD4 mitochondrial localization rescues respiratory function in AIF-deficient cells and enables programmed cell death for embryonic morphogenesis. Co-immunoprecipitation, siRNA knockdown, rescue by forced mitochondrial localization of CHCHD4, embryoid body cavitation assay Molecular cell High 26004228
2015 Mia40 (CHCHD4) interacts with and introduces an intermolecular disulfide bond that links MICU1 and MICU2 into a heterodimer; this disulfide-dependent heterodimer associates with MCU at low Ca2+ and dissociates at high Ca2+, thereby regulating mitochondrial Ca2+ uptake. Mia40 interactome (MS), in vitro disulfide bond formation assays, Co-IP, Ca2+ uptake measurements, mutagenesis Cell metabolism High 26387864
2015 AIF physically interacts with CHCHD4/MIA40 in patient fibroblasts and mouse tissues; AIF deficiency correlates with decreased MIA40 protein levels without changes in mRNA; MIA40 overexpression counteracts loss of respiratory subunits in AIF-deficient (Harlequin) cells. Co-immunoprecipitation, western blot, MIA40 overexpression rescue in Hq mouse cells and patient fibroblasts Cell death & disease High 26158520
2015 Mia40 functions as an electron sink: it can accept up to six electrons from substrates (rather than just two), resulting in fully reduced Mia40 that undergoes conformational change; this enables insertion of two disulfide bonds per substrate without requiring a ternary Erv1-Mia40-substrate complex. In vitro oxidation assays with Tim13, electron counting by redox state analysis, limited proteolysis of reduced Mia40, erv1-101 in organello trapping The Journal of biological chemistry High 26085103
2016 Mia40's substrate-binding domain (hydrophobic cleft) is both necessary and sufficient to promote protein import (trapping substrates as a 'holding trap'), while the CPC oxidase domain is required for viability but can be partially replaced by the chemical oxidant diamide; this demonstrates that Mia40 acts primarily as a trans-site receptor driving translocation via hydrophobic substrate binding. Domain-dissection mutagenesis, yeast complementation, diamide rescue assay, import assays eLife High 27343349
2016 Exercise downregulates CHCHD4 via FOXO3 binding to the CHCHD4 promoter, which reduces p53 import into mitochondria and increases nuclear p53 localization; nuclear p53 and FOXO3 then synergistically transactivate SIRT1; transgenic mice with constitutive CHCHD4 expression lose this exercise-induced response. ChIP (FOXO3 on CHCHD4 promoter), transgenic mouse model, p53 localization assays, luciferase reporter assays The Journal of biological chemistry Medium 27687729
2017 Elevated CHCHD4 expression or prolonged hypoxia causes perinuclear accumulation of mitochondria in a HIF-1α-dependent manner; CHCHD4 is required for perinuclear mitochondrial localization and HIF activation in hypoxia; mutation of the CPC motif cysteines or inhibition of complex IV redistributes mitochondria peripherally and blocks HIF activation. Live imaging of mitochondrial distribution, CPC mutagenesis, HIF-1α siRNA, complex IV inhibition (sodium azide), CHCHD4 overexpression/knockdown Frontiers in oncology Medium 28497026
2018 Human CHCHD4 exists in a predominantly oxidized state in vivo (70–90% oxidized in mouse tissues and cultured cells); ALR is superstoichiometric over CHCHD4 in most tissues but does not fully oxidize CHCHD4, indicating regulated redox balance. In vivo redox state measurement (thiol-trapping/alkylation + mass spectrometry), quantitative proteomics across tissues Redox biology Medium 29704824
2018 pVHL re-expression in pVHL-defective renal carcinoma cells elevates CHCHD4 expression alongside respiratory chain subunits (NDUFB10, mtCO-2, COX IV), increased oxygen consumption, and altered metabolism; knockdown of HIF-2α similarly elevates CHCHD4; this establishes a VHL/HIF-2α axis regulating CHCHD4 and mitochondrial function. pVHL re-expression, HIF-2α siRNA knockdown, SILAC proteomics, OCR measurement Frontiers in oncology Medium 30338240
2018 CHCHD10 (a CHCH domain protein linked to ALS) requires Mia40 for its mitochondrial import via its CHCH domain (not the N-terminal targeting sequence); ALS-associated mutation Q108P nearly completely blocks import; Mia40 overexpression rescues import of CHCHD10 Q108P by enhancing disulfide bond formation. Mia40 knockdown/overexpression, CHCHD10 truncation analysis, import assays, patient mutation analysis EMBO molecular medicine Medium 29789341
2020 Human MIA40 (CHCHD4) undergoes reversible S-glutathionylation at three cysteines in the twin CX9C motifs and Cys4; cells expressing glutathionylation-deficient MIA40 show reduced complex III and IV activities and elevated ROS; glutathionylated MIA40 can directly transfer electrons to cytochrome c, with Fe-S clusters in the CPC motif essential for the two-electron to one-electron transfer. Site-directed mutagenesis, MALDI mass spectrometry of in vivo protein, immunocapture of complex III, electron transfer assays Redox biology Medium 32971361
2020 The conserved negatively charged C-terminal region of human MIA40 (CHCHD4) is dispensable for redox function but critical during cytosolic residence before import: it slows the import half-time (~90 min) and protects the unfolded cytosolic MIA40 precursor from proteasomal degradation; this stabilizing function is transferable to another IMS precursor (COX19). C-terminal deletion/mutagenesis, cycloheximide chase, proteasome inhibitor treatment, import kinetics assays BMC biology Medium 32762682
2021 Increased levels of Mia40 (CHCHD4) suppress formation of toxic polyQ (Q97-GFP) aggregates in yeast and human cells by competing for chaperone and proteasome capacity; Mia40 has a rate-limiting role in mitochondrial protein import that regulates cytosolic proteostasis. Mia40 overexpression in yeast and human cells, polyQ aggregation assays, cell viability assays, mia40 mutant hypersensitivity The EMBO journal Medium 34191328
2023 CHCHD4 controls TRIAP1 mitochondrial import; exercise-induced downregulation of CHCHD4 (via FOXO3) decreases TRIAP1 import into mitochondria, reducing cardiolipin levels and promoting VDAC oligomerization, which facilitates mtDNA release, activating cGAS-STING/NF-κB innate immune signaling that drives slow-twitch muscle fiber formation; CHCHD4 haploinsufficiency phenocopies this in mice. CHCHD4 haploinsufficient mice, VDAC oligomerization assays, cardiolipin measurement, cGAS-STING pathway activation, TRIAP1 import assays Cell reports Medium 38157298
2024 Crystal structure of the ternary complex of AIF with NAD+, FAD, and the N-terminal 27-mer peptide of CHCHD4 reveals the structural basis of AIF-CHCHD4 interaction: the CHCHD4 N-terminal peptide binds at the AIF active site and exerts allosteric effects on the cofactor-binding site; validated by cross-linking mass spectrometry and site-directed mutagenesis. X-ray crystallography, chemical cross-linking mass spectrometry, site-directed mutagenesis, biophysical binding assays Structure (London, England : 1993) High 38460521
2024 MIA40 (CHCHD4) non-covalently interacts with the cysteine-free IMS protein HAX1 via its holdase (hydrophobic cleft) activity rather than oxidoreductase activity; this interaction stabilizes HAX1 in the IMS and prevents its aggregation and degradation; loss of MIA40 leads to HAX1 aggregation, identifying a cysteine-independent substrate requiring only MIA40 chaperone activity. Co-immunoprecipitation, in vitro binding assays, MIA40 KO/KD, redox-active cysteine mutagenesis of MIA40 The FEBS journal Medium 39564806
2025 MIA40 (CHCHD4) accelerates TRIAP1 oxidative folding 30-fold by bypassing a non-native Cys37-Cys47 kinetic trap; MIA40 drives formation of the inner disulfide (Cys18-Cys37) first, then can catalyze the outer disulfide (Cys8-Cys47); TRIAP1's reduced state in the cytoplasm is a functional molten globule linked to its cytosolic apoptosis-inhibitory role. In vitro oxidative folding kinetics, NMR characterization of folding intermediates, disulfide mapping by mass spectrometry The Journal of biological chemistry High 39909379
2025 MIA40 (CHCHD4) forms a complex with AIFM1 in the IMS that suppresses AIFM1-induced cell death in a NADH-dependent manner; increased NADH/NAD+ ratio (due to complex I dysfunction) strengthens the AIFM1-MIA40 interaction; MIA40 silencing, complex I rescue, or NADH depletion sensitizes complex I-deficient cells to AIFM1-mediated apoptosis. Complex I KO cell lines (NDUFA13-KO), MIA40 siRNA, NADH depletion by yeast NADH oxidase expression, cell death assays, Co-IP EMBO reports Medium 40055465
2025 FAM136A is a new MIA40 substrate in the IMS: MIA40 introduces four disulfide bonds in two twin-CX3C motifs of FAM136A; FAM136A steady-state levels are strongly dependent on MIA40 and AIFM1 levels; loss of FAM136A triggers integrated stress response and causes aggregation of other IMS proteins (HAX1, CLPB). In vitro disulfide bond formation assays, MIA40/AIFM1 knockdown, co-immunoprecipitation, acute genetic deletion, proteomics bioRxivpreprint Medium bio_10.1101_2025.09.22.677734

Source papers

Stage 0 corpus · 80 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2004 Essential role of Mia40 in import and assembly of mitochondrial intermembrane space proteins. The EMBO journal 367 15359280
2009 MIA40 is an oxidoreductase that catalyzes oxidative protein folding in mitochondria. Nature structural & molecular biology 220 19182799
2005 Erv1 mediates the Mia40-dependent protein import pathway and provides a functional link to the respiratory chain by shuttling electrons to cytochrome c. Journal of molecular biology 198 16185707
2015 Interaction between AIF and CHCHD4 Regulates Respiratory Chain Biogenesis. Molecular cell 166 26004228
2015 The Ca(2+)-Dependent Release of the Mia40-Induced MICU1-MICU2 Dimer from MCU Regulates Mitochondrial Ca(2+) Uptake. Cell metabolism 164 26387864
2009 A novel intermembrane space-targeting signal docks cysteines onto Mia40 during mitochondrial oxidative folding. The Journal of cell biology 153 20026652
2005 Mia40, a novel factor for protein import into the intermembrane space of mitochondria is able to bind metal ions. FEBS letters 144 15620710
2005 The essential mitochondrial protein Erv1 cooperates with Mia40 in biogenesis of intermembrane space proteins. Journal of molecular biology 130 16181637
2010 Molecular chaperone function of Mia40 triggers consecutive induced folding steps of the substrate in mitochondrial protein import. Proceedings of the National Academy of Sciences of the United States of America 118 21059946
2005 Functional and mutational characterization of human MIA40 acting during import into the mitochondrial intermembrane space. Journal of molecular biology 99 16185709
2011 Molecular recognition and substrate mimicry drive the electron-transfer process between MIA40 and ALR. Proceedings of the National Academy of Sciences of the United States of America 95 21383138
2007 The Erv1-Mia40 disulfide relay system in the intermembrane space of mitochondria. Biochimica et biophysica acta 95 18179776
2010 Conserved and novel functions for Arabidopsis thaliana MIA40 in assembly of proteins in mitochondria and peroxisomes. The Journal of biological chemistry 92 20829360
2009 Structural basis of yeast Tim40/Mia40 as an oxidative translocator in the mitochondrial intermembrane space. Proceedings of the National Academy of Sciences of the United States of America 91 19667201
2012 Human CHCHD4 mitochondrial proteins regulate cellular oxygen consumption rate and metabolism and provide a critical role in hypoxia signaling and tumor progression. The Journal of clinical investigation 83 22214851
2015 Loss of apoptosis-inducing factor critically affects MIA40 function. Cell death & disease 82 26158520
2012 Atp23 biogenesis reveals a chaperone-like folding activity of Mia40 in the IMS of mitochondria. The EMBO journal 82 22990235
2007 Oxidative folding of small Tims is mediated by site-specific docking onto Mia40 in the mitochondrial intermembrane space. Molecular microbiology 78 17680986
2017 Repurposing Approach Identifies Auranofin with Broad Spectrum Antifungal Activity That Targets Mia40-Erv1 Pathway. Frontiers in cellular and infection microbiology 75 28149831
2007 Biogenesis of the essential Tim9-Tim10 chaperone complex of mitochondria: site-specific recognition of cysteine residues by the intermembrane space receptor Mia40. The Journal of biological chemistry 73 17553782
2013 Mitochondrial protein import: Mia40 facilitates Tim22 translocation into the inner membrane of mitochondria. Molecular biology of the cell 70 23283984
2011 Anamorsin is a [2Fe-2S] cluster-containing substrate of the Mia40-dependent mitochondrial protein trapping machinery. Chemistry & biology 65 21700214
2008 Structural and functional roles of the conserved cysteine residues of the redox-regulated import receptor Mia40 in the intermembrane space of mitochondria. The Journal of biological chemistry 65 19011240
2016 Mia40 is a trans-site receptor that drives protein import into the mitochondrial intermembrane space by hydrophobic substrate binding. eLife 64 27343349
2009 Reconstitution of the mia40-erv1 oxidative folding pathway for the small tim proteins. Molecular biology of the cell 63 19477928
2008 The zinc-binding protein Hot13 promotes oxidation of the mitochondrial import receptor Mia40. EMBO reports 63 18787558
2011 Mia40-dependent oxidation of cysteines in domain I of Ccs1 controls its distribution between mitochondria and the cytosol. Molecular biology of the cell 56 21865594
2008 Mitochondrial biogenesis, switching the sorting pathway of the intermembrane space receptor Mia40. The Journal of biological chemistry 56 18779329
2007 The sulfhydryl oxidase Erv1 is a substrate of the Mia40-dependent protein translocation pathway. FEBS letters 55 17336303
2014 Mia40 targets cysteines in a hydrophobic environment to direct oxidative protein folding in the mitochondria. Nature communications 52 24407114
2012 Disulfide bond formation: sulfhydryl oxidase ALR controls mitochondrial biogenesis of human MIA40. Traffic (Copenhagen, Denmark) 46 23186364
2018 A novel CHCHD10 mutation implicates a Mia40-dependent mitochondrial import deficit in ALS. EMBO molecular medicine 45 29789341
2020 AIF meets the CHCHD4/Mia40-dependent mitochondrial import pathway. Biochimica et biophysica acta. Molecular basis of disease 44 32105825
2012 In vivo evidence for cooperation of Mia40 and Erv1 in the oxidation of mitochondrial proteins. Molecular biology of the cell 44 22918950
2013 Mia40 and MINOS act in parallel with Ccs1 in the biogenesis of mitochondrial Sod1. The FEBS journal 36 23802566
2010 The N-terminal shuttle domain of Erv1 determines the affinity for Mia40 and mediates electron transfer to the catalytic Erv1 core in yeast mitochondria. Antioxidants & redox signaling 32 20367271
2017 CHCHD4 Regulates Intracellular Oxygenation and Perinuclear Distribution of Mitochondria. Frontiers in oncology 30 28497026
2021 Increased levels of mitochondrial import factor Mia40 prevent the aggregation of polyQ proteins in the cytosol. The EMBO journal 27 34191328
2021 CHCHD4 (MIA40) and the mitochondrial disulfide relay system. Biochemical Society transactions 24 33599699
2021 The Mia40/CHCHD4 Oxidative Folding System: Redox Regulation and Signaling in the Mitochondrial Intermembrane Space. Antioxidants (Basel, Switzerland) 24 33921425
2017 Erv1 of Arabidopsis thaliana can directly oxidize mitochondrial intermembrane space proteins in the absence of redox-active Mia40. BMC biology 24 29117860
2018 VHL-Mediated Regulation of CHCHD4 and Mitochondrial Function. Frontiers in oncology 23 30338240
2013 Identification and characterization of mitochondrial Mia40 as an iron-sulfur protein. The Biochemical journal 23 23834247
2019 CHCHD4 regulates tumour proliferation and EMT-related phenotypes, through respiratory chain-mediated metabolism. Cancer & metabolism 21 31346464
2017 Haploinsufficiency in the mitochondrial protein CHCHD4 reduces brain injury in a mouse model of neonatal hypoxia-ischemia. Cell death & disease 21 28492551
2020 The C-terminal region of the oxidoreductase MIA40 stabilizes its cytosolic precursor during mitochondrial import. BMC biology 20 32762682
2018 The mitochondrial oxidoreductase CHCHD4 is present in a semi-oxidized state in vivo. Redox biology 20 29704824
2014 Mia40 is optimized for function in mitochondrial oxidative protein folding and import. ACS chemical biology 20 24983157
2014 Mia40 combines thiol oxidase and disulfide isomerase activity to efficiently catalyze oxidative folding in mitochondria. Journal of molecular biology 20 25451030
2016 Activation of Mitochondrial Protein Phosphatase SLP2 by MIA40 Regulates Seed Germination. Plant physiology 19 27923987
2015 Cytosolic Fe-S Cluster Protein Maturation and Iron Regulation Are Independent of the Mitochondrial Erv1/Mia40 Import System. The Journal of biological chemistry 19 26396185
2013 Biogenesis of yeast Mia40 - uncoupling folding from import and atypical recognition features. The FEBS journal 19 23937629
2020 Glutathionylated and Fe-S cluster containing hMIA40 (CHCHD4) regulates ROS and mitochondrial complex III and IV activities of the electron transport chain. Redox biology 18 32971361
2019 CHCHD4 confers metabolic vulnerabilities to tumour cells through its control of the mitochondrial respiratory chain. Cancer & metabolism 17 30886710
2016 Forkhead Box O3A (FOXO3) and the Mitochondrial Disulfide Relay Carrier (CHCHD4) Regulate p53 Protein Nuclear Activity in Response to Exercise. The Journal of biological chemistry 16 27687729
2014 The mitochondrial intermembrane space oxireductase Mia40 funnels the oxidative folding pathway of the cytochrome c oxidase assembly protein Cox19. The Journal of biological chemistry 16 24569988
2023 Impaired AIF-CHCHD4 interaction and mitochondrial calcium overload contribute to auditory neuropathy spectrum disorder in patient-iPSC-derived neurons with AIFM1 variant. Cell death & disease 14 37365177
2019 Santacruzamate A Ameliorates AD-Like Pathology by Enhancing ER Stress Tolerance Through Regulating the Functions of KDELR and Mia40-ALR in vivo and in vitro. Frontiers in cellular neuroscience 14 30886573
2015 Mia40 Protein Serves as an Electron Sink in the Mia40-Erv1 Import Pathway. The Journal of biological chemistry 14 26085103
2015 CHCHD4 links AIF to the biogenesis of respiratory chain complex I. Molecular & cellular oncology 14 27308594
2015 Human mitochondrial MIA40 (CHCHD4) is a component of the Fe-S cluster export machinery. The Biochemical journal 13 26275620
2015 Mia40 is a facile oxidant of unfolded reduced proteins but shows minimal isomerase activity. Archives of biochemistry and biophysics 12 26014136
2015 Metabolic epistasis among apoptosis-inducing factor and the mitochondrial import factor CHCHD4. Cell cycle (Georgetown, Tex.) 12 26178476
2023 CHCHD4-TRIAP1 regulation of innate immune signaling mediates skeletal muscle adaptation to exercise. Cell reports 11 38157298
2017 A single-cysteine mutant and chimeras of essential Leishmania Erv can complement the loss of Erv1 but not of Mia40 in yeast. Redox biology 10 29310075
2024 CHCHD4 binding affects the active site of apoptosis inducing factor (AIF): Structural determinants for allosteric regulation. Structure (London, England : 1993) 5 38460521
2023 Elevated CHCHD4 orchestrates mitochondrial oxidative phosphorylation to disturb hypoxic pulmonary hypertension. Journal of translational medicine 5 37438854
2024 CHCHD4 regulates the expression of mitochondrial genes that are essential for tumour cell growth. Biochimica et biophysica acta. Molecular basis of disease 4 38909850
2024 Interaction with the cysteine-free protein HAX1 expands the substrate specificity and function of MIA40 beyond protein oxidation. The FEBS journal 4 39564806
2022 USF1-CHCHD4 axis promotes lung adenocarcinoma progression partially via activating the MYC pathway. Discover oncology 4 36482116
2021 Moschamindole induces glioma cell apoptosis by blocking Mia40-dependent mitochondrial intermembrane space assembly and oxidative respiration. Phytotherapy research : PTR 4 33856743
2025 MIA40 circumvents the folding constraints imposed by TRIAP1 function. The Journal of biological chemistry 3 39909379
2018 Proteasomal degradation competes with Mia40-mediated import into mitochondria. BMC biology 3 29929505
2025 CHCHD4 Oxidoreductase Activity: A Comprehensive Analysis of the Molecular, Functional, and Structural Properties of Its Redox-Regulated Substrates. Molecules (Basel, Switzerland) 2 40430290
2024 [Mechanism of Liangfang Wenjing Decoction in treatment of hypoxia on endometriosis with cold coagulation and blood stasis by regulating CHCHD4 expression]. Zhongguo Zhong yao za zhi = Zhongguo zhongyao zazhi = China journal of Chinese materia medica 2 39099355
2025 MIA40 suppresses cell death induced by apoptosis-inducing factor 1. EMBO reports 1 40055465
2026 Biallelic variants in CHCHD4 are associated with combined OXPHOS defect leading to mitochondrial disease. HGG advances 0 41981912
2025 The Mia40 substrate Mix17 exposes its N-terminus to the cytosolic side of the mitochondrial outer membrane. Journal of cell science 0 40094392
2025 RETRACTION: Moschamindole Induces Glioma Cell Apoptosis by Blocking Mia40-Dependent Mitochondrial Intermembrane Space Assembly and Oxidative Respiration. Phytotherapy research : PTR 0 40331711
2021 Corrigendum: VHL-Mediated Regulation of CHCHD4 and Mitochondrial Function. Frontiers in oncology 0 34631576