Affinage

CYB5A

Cytochrome b5 · UniProt P00167

Length
134 aa
Mass
15.3 kDa
Annotated
2026-04-28
26 papers in source corpus 11 papers cited in narrative 11 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

CYB5A (microsomal cytochrome b5) is an electron-transfer cofactor central to androgen biosynthesis and cholesterol pathway regulation. It stimulates CYP17A1 17,20-lyase activity to drive production of DHEA, androstenedione, and 16-androstene steroids, with specific surface residues (including N62, N21, L28, R52) mediating the CYB5A–CYP17A1 interaction (PMID:31509771, PMID:21185375, PMID:25890314); homozygous loss-of-function mutations in CYB5A cause isolated 17,20-lyase deficiency with methemoglobinemia in humans (PMID:32051920). CYB5A and its mitochondrial paralog CYB5B compensate for each other in sterol C4-demethylation during cholesterol biosynthesis, and their combined loss causes accumulation of dimethyl sterol intermediates (T-MAS), suppressing the SREBP pathway and activating PPARγ signaling (PMID:39489939). CYB5A also promotes autophagy through the AKT/mTOR/ULK1 axis, a function linked to osteogenic differentiation and suppression of oncogenic phenotypes in pancreatic cancer (PMID:40246926, PMID:24448000).

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 2005 Medium

    A naturally occurring 5′ UTR polymorphism in porcine CYB5A directly reduced CYB5A enzymatic activity and lowered androstenone levels in fat, establishing CYB5A as a functional determinant of 16-androstene steroid biosynthesis.

    Evidence Functional activity assay comparing CYB5A activity between genotypes in 229 porcine testis samples

    PMID:16104384

    Open questions at the time
    • Mechanism by which CYB5A stimulates 16-androstene synthesis not defined at residue level
    • No human data provided
  2. 2010 Medium

    siRNA knockdown in human adrenal cells demonstrated that CYB5A is required for androstenedione production, connecting CYB5A's cofactor role specifically to human adrenal androgen output.

    Evidence siRNA knockdown in H295R cells with androstenedione measurement; co-immunolocalization in human adrenal cortex

    PMID:21185375

    Open questions at the time
    • Whether CYB5A directly transfers electrons to CYP17A1 versus acts allosterically was not resolved
    • Single cell line system
  3. 2014 Medium

    Promoter SNPs in porcine CYB5A were shown to alter transcription factor (MAZ) binding and expression levels, revealing transcriptional regulatory mechanisms controlling CYB5A abundance.

    Evidence EMSA and luciferase reporter assays in transfected cells with quantitative PCR from liver tissues

    PMID:25516134

    Open questions at the time
    • Whether MAZ-dependent regulation operates in human CYB5A promoter unknown
    • Downstream metabolic consequences of expression differences not measured
  4. 2014 Low

    CYB5A was linked to autophagy induction and tumor suppression in pancreatic cancer, broadening its known functions beyond electron transfer to include signaling through TRAF6.

    Evidence In vitro and in vivo loss-of-function studies with autophagy readouts and TRAF6 pathway analysis

    PMID:24448000

    Open questions at the time
    • Limited mechanistic detail; no reconstitution or mutagenesis to establish direct CYB5A–TRAF6 interaction
    • Single lab with no independent confirmation
    • How an ER-anchored electron carrier triggers autophagy is unexplained
  5. 2015 Medium

    A human intronic CYB5A SNP that doubles mRNA expression was shown to produce 2–3 fold activation of CYP17A1 17,20-lyase activity, quantitatively linking CYB5A expression levels to androgen synthetic capacity.

    Evidence Radiolabeled steroid conversion assays in synovial fibroblasts; qRT-PCR; immunohistochemistry

    PMID:25890314

    Open questions at the time
    • Studied in synovial fibroblasts rather than adrenal or gonadal tissue
    • Allosteric versus electron-transfer mechanism not distinguished
  6. 2019 High

    Site-directed mutagenesis of specific CYB5A surface residues (N62S, N21K, L28V, R52M) altered DHEA and 16-androstene production when co-expressed with CYP17A1, identifying the molecular interface that governs CYB5A's stimulation of 17,20-lyase activity.

    Evidence Mutagenesis of CYB5A and CYP17A1 expressed in HEK293 cells; radiolabeled pregnenolone conversion; HPLC metabolite quantification

    PMID:31509771

    Open questions at the time
    • No crystal structure of the CYB5A–CYP17A1 complex to confirm binding mode
    • Whether the same residues are critical in human versus porcine CYB5A not tested
  7. 2019 Medium

    A homozygous nonsense CYB5A mutation (p.Tyr35Ter) in a human patient caused isolated 17,20-lyase deficiency with methemoglobinemia, providing in vivo genetic proof that CYB5A is essential for CYP17A1 lyase activity and methemoglobin reduction.

    Evidence Human genetic case study with hormonal profiling and DNA sequencing

    PMID:32051920

    Open questions at the time
    • Single family; full phenotypic spectrum of CYB5A deficiency not delineated
    • No functional rescue experiment performed
  8. 2022 Medium

    An alternative CYB5A transcript (CYB5Aalt) expressed in aneuploid B-ALL was found to downregulate canonical CYB5A functions and increase resistance to BCL2-mediated apoptosis, revealing that alternative CYB5A isoforms can act as dominant-negative regulators.

    Evidence RNA-seq discovery; stable CYB5Aalt overexpression in NALM-6 cells with transcriptomic and cell death assays

    PMID:35436854

    Open questions at the time
    • Protein product of CYB5Aalt not biochemically characterized
    • Whether CYB5Aalt is recurrently expressed in other cancers unknown
  9. 2024 High

    Combined loss of CYB5A and CYB5B was shown to block sterol C4-demethylation, causing T-MAS accumulation that suppresses SREBP and activates PPARγ, establishing functional redundancy between the two cytochrome b5 paralogs in cholesterol biosynthesis.

    Evidence CRISPR knockout in HeLa cells; liver-specific Cyb5b KO mice with shRNA knockdown of Cyb5a; sterol profiling and pathway analysis

    PMID:39489939

    Open questions at the time
    • Which C4-demethylation enzyme(s) CYB5A directly donates electrons to was not identified
    • Tissue-specific relative contributions of CYB5A vs CYB5B not fully resolved
  10. 2025 Medium

    CYB5A was shown to promote osteogenic differentiation by activating autophagy through the AKT/mTOR/ULK1 axis, providing a defined signaling pathway for CYB5A's autophagy-stimulating function first noted in cancer cells.

    Evidence Overexpression and siRNA knockdown in MC3T3-E1 cells; Western blotting for autophagy markers; pharmacological autophagy modulators

    PMID:40246926

    Open questions at the time
    • How an ER-membrane electron carrier engages AKT/mTOR signaling is mechanistically unexplained
    • Single cell line (osteoblast precursor); generalizability unclear

Open questions

Synthesis pass · forward-looking unresolved questions
  • The structural basis of CYB5A interaction with CYP17A1 and the C4-demethylation machinery, the mechanism by which CYB5A activates autophagy signaling, and the physiological significance of the alternative CYB5Aalt isoform remain unresolved.
  • No crystal or cryo-EM structure of CYB5A–CYP17A1 complex
  • Electron-transfer versus allosteric mechanism for 17,20-lyase stimulation not resolved
  • Molecular link between CYB5A and AKT/mTOR unknown

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016491 oxidoreductase activity 4 GO:0140104 molecular carrier activity 2
Localization
GO:0005783 endoplasmic reticulum 2
Pathway
R-HSA-1430728 Metabolism 5 R-HSA-9612973 Autophagy 2
Partners
CYB5BCYP17A1HSD3B2TRAF6

Evidence

Reading pass · 11 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2010 CYB5A is required for androstenedione production in human adrenal H295R cells; siRNA knockdown of CYB5A significantly inhibited androstenedione production, and CYB5A co-localizes with HSD3B2 in hybrid ZF/ZR border cells of the human adrenal cortex. siRNA knockdown in H295R cells with androstenedione measurement; co-immunolocalization in human adrenal tissue The Journal of steroid biochemistry and molecular biology Medium 21185375
2015 A CYB5A intronic SNP (rs1790834) doubles CYB5A mRNA expression and results in a 2–3 fold activation of steroid 17,20-lyase (CYP17A1) activity, demonstrating that CYB5A is a critical cofactor for 17,20-lyase activity and androgen synthesis. Radiolabeled steroid conversion assays in synovial fibroblasts; quantitative RT-PCR; immunohistochemistry; GWAS association study Arthritis research & therapy Medium 25890314
2005 A SNP in the 5' UTR of porcine CYB5A (G→T at -8 upstream of ATG) reduces CYB5A enzymatic activity and is associated with lower androstenone levels in fat, linking CYB5A activity directly to 16-androstene (androstenone) biosynthesis. Functional activity assay comparing CYB5A activity between genotypes; genotyping of 229 testis samples Mammalian genome Medium 16104384
2019 Specific surface residues of porcine CYB5A are involved in binding to CYP17A1; mutations at these residues (including N62S, N21K, L28V, R52M) alter the production of DHEA and 16-androstene steroids, with N62S increasing both 16A and DHEA production when combined with wild-type CYP17A1. Site-directed mutagenesis of CYB5A and CYP17A1; expression in HEK293 cells with radiolabeled pregnenolone substrate; HPLC metabolite quantification The Journal of steroid biochemistry and molecular biology High 31509771
2019 A homozygous loss-of-function CYB5A mutation (p.Tyr35Ter) causes isolated 17,20-lyase deficiency with methemoglobinemia in humans, confirming that CYB5A is required for CYP17A1 17,20-lyase activity in vivo. Human genetic case study; hormonal studies confirming isolated 17,20-lyase deficiency; DNA sequencing Journal of the Endocrine Society Medium 32051920
2024 CYB5A (microsomal) and CYB5B (mitochondrial) compensate for each other in sterol C4-demethylation during cholesterol biosynthesis; knockout of CYB5B alone in HeLa cells blocks cholesterol biosynthesis causing T-MAS accumulation, and combined knockdown of both CYB5A and CYB5B in liver-specific Cyb5b KO mice leads to marked T-MAS/dihydro-T-MAS accumulation, inhibiting the SREBP pathway and activating the PPARγ pathway. CRISPR knockout in HeLa cells; liver-specific Cyb5b KO mice with shRNA knockdown of Cyb5a; sterol profiling; SREBP and PPARγ pathway analysis Cell reports High 39489939
2014 CYB5A inhibits oncogenic phenotypes in pancreatic cancer cells through induction of autophagy, and its downstream pathway involves TRAF6. In vitro and in vivo studies with CYB5A loss-of-function; autophagy readouts; pathway analysis involving TRAF6 Autophagy Low 24448000
2025 CYB5A promotes osteogenic differentiation of MC3T3-E1 cells by stimulating autophagy via activation of ULK1 and reduction of mTOR phosphorylation (AKT/mTOR/ULK1 pathway); CYB5A overexpression increases LC3-II/LC3-I ratio and decreases P62, and autophagy inhibitors/activators confirmed this pathway mediates CYB5A's effect on osteogenesis. Overexpression and siRNA knockdown in MC3T3-E1 cells; Western blotting for LC3-II/I, P62, mTOR phosphorylation, ULK1; autophagy inhibitor/activator pharmacological validation Scientific reports Medium 40246926
2012 CYB5A (cytochrome b5) functions to detoxify sulfonamide hydroxylamine metabolites, with CYB5A activity serving as part of a reductive detoxification pathway for aromatic amine intermediates. Candidate gene sequencing; genotyping of CYB5A polymorphisms in sulfonamide hypersensitivity cases vs controls Pharmacogenetics and genomics Low 22850190
2014 Promoter SNPs in porcine CYB5A affect transcription factor (MAZ) binding and alter CYB5A expression levels, as demonstrated by EMSA and reporter assays, with haplotype B showing higher CYB5A expression than haplotype A. Electrophoretic mobility shift assay (EMSA); in vitro cell transfection with luciferase reporter assay; quantitative PCR from liver tissues; in silico transcription factor prediction Animal genetics Medium 25516134
2022 An alternative CYB5A transcript (CYB5Aalt) with a novel transcription start site and non-canonical exon usage is expressed in aneuploid B-cell precursor ALL; overexpression of CYB5Aalt in NALM-6 cells downregulates pathways related to wild-type CYB5A functions, decreases wild-type CYB5A expression, and increases resistance to BCL2-mediated apoptosis. RNA sequencing; stable overexpression of CYB5Aalt in NALM-6 cells; differential expression and GSEA analysis; cell death/viability assays BMC genomic data Medium 35436854

Source papers

Stage 0 corpus · 26 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2010 Human adrenal cells that express both 3β-hydroxysteroid dehydrogenase type 2 (HSD3B2) and cytochrome b5 (CYB5A) contribute to adrenal androstenedione production. The Journal of steroid biochemistry and molecular biology 28 21185375
2015 CYB5A polymorphism increases androgens and reduces risk of rheumatoid arthritis in women. Arthritis research & therapy 24 25890314
2014 3βHSD and CYB5A double positive adrenocortical cells during adrenal development/aging. Endocrine research 20 24832628
2005 A novel polymorphism in the 5' untranslated region of the porcine cytochrome b5 (CYB5) gene is associated with decreased fat androstenone level. Mammalian genome : official journal of the International Mammalian Genome Society 20 16104384
2012 Evaluation of polymorphisms in the sulfonamide detoxification genes NAT2, CYB5A, and CYB5R3 in patients with sulfonamide hypersensitivity. Pharmacogenetics and genomics 15 22850190
2014 Polymorphisms in the carcinogen detoxification genes CYB5A and CYB5R3 and breast cancer risk in African American women. Cancer causes & control : CCC 13 25225034
2014 Differential expression of CYB5A in Chinese and European pig breeds due to genetic variations in the promoter region. Animal genetics 13 25516134
2014 CYB5A and autophagy-mediated cell death in pancreatic cancer. Autophagy 12 24448000
2019 Isolated 17,20-Lyase Deficiency in a CYB5A Mutated Female With Normal Sexual Development and Fertility. Journal of the Endocrine Society 11 32051920
2004 Disruption of the Candida albicans CYB5 gene results in increased azole sensitivity. Antimicrobial agents and chemotherapy 10 15328107
2019 Effect of mutations in porcine CYB5A and CYP17A1 on the metabolism of pregnenolone. The Journal of steroid biochemistry and molecular biology 8 31509771
2012 Evaluation of polymorphisms in the sulfonamide detoxification genes CYB5A and CYB5R3 in dogs with sulfonamide hypersensitivity. Journal of veterinary internal medicine 7 22816446
2022 Functional variants in cytochrome b5 type A (CYB5A) are enriched in Southwest American Indian individuals and associate with obesity. Obesity (Silver Spring, Md.) 6 35043601
2016 2,3,7,8 Tetrachlorodibenzo-p-dioxin-induced RNA abundance changes identify Ackr3, Col18a1, Cyb5a and Glud1 as candidate mediators of toxicity. Archives of toxicology 6 27136898
2008 Effect of polymorphism in the porcine cytochrome b5 ( CYB5A) gene on androstenone and skatole concentrations and sexual development in Swedish pig populations. Animal : an international journal of animal bioscience 6 22445012
2025 CYB5A promotes osteogenic differentiation of MC3T3-E1 cells through autophagy mediated by the AKT/mTOR/ULK1 signaling pathway. Scientific reports 5 40246926
2024 Defects in CYB5A and CYB5B impact sterol-C4 oxidation in cholesterol biosynthesis and demonstrate regulatory roles of dimethyl sterols. Cell reports 5 39489939
2021 Isolated 17, 20 Lyase Deficiency Secondary to a Novel CYB5A Variant: Comparison of Steroid Metabolomic Findings with Published Cases Provides Diagnostic Guidelines and Greater Insight into Its Biological Role. Hormone research in paediatrics 4 33626548
2021 Lack of association between CYB5A gene rs1790834 polymorphism and the response to leflunomide in women with rheumatoid arthritis. European journal of clinical pharmacology 3 34160668
2025 Two families, two pathways: a case series of 46, XY DSD with 17α-hydroxylase deficiency and isolated 17,20-lyase deficiency due to novel CYB5A variant. Journal of pediatric endocrinology & metabolism : JPEM 2 40123165
2023 The association between CYB5A gene rs1790834 polymorphism and clinical improvement after 6 months of leflunomide treatment in women with rheumatoid arthritis. Clinical rheumatology 1 37289314
2022 An alternative CYB5A transcript is expressed in aneuploid ALL and enriched in relapse. BMC genomic data 1 35436854
2016 Development of CYB5-fusion monitoring system for efficient periplasmic expression of multimeric proteins in Escherichia coli. Protein expression and purification 1 27524697
2026 The azole-resistance phenotype of a Nakaseomyces glabratus clinical strain encoding a wild-type PDR1 allele involves the efflux pumps Aus1 and Pdh1 and Cyb5, an alternative reductase required for ergosterol biosynthesis. Microbiology spectrum 0 41891695
2025 Haplotype-based association of CYB5A gene polymorphisms (rs1790834 and rs1790858) with polycystic ovary syndrome in a south Indian cohort. Gene 0 41052660
2025 Micro RNA miR-726-3p targets CYB5A in Hen ovaries to modulate granulosa cell proliferation and differentiation. Scientific reports 0 41453937