Affinage

MID2

Probable E3 ubiquitin-protein ligase MID2 · UniProt Q9UJV3

Length
735 aa
Mass
83.2 kDa
Annotated
2026-06-10
30 papers in source corpus 13 papers cited in narrative 14 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 7/7 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

MID2 (TRIM1/FXY2) is a microtubule-associated RING-domain E3 ubiquitin ligase that couples substrate ubiquitination to the cytoskeleton to control mitotic progression, kinase regulation, and signaling (PMID:26748699, PMID:35266954). It localizes to microtubules through its C-terminal B30.2 domain and its COS domain, the latter being indispensable for cytoskeletal targeting since a COS-domain missense mutation (p.Arg347Gln) redistributes the protein to the cytoplasm (PMID:10644436, PMID:24115387); MID2 also homo- and heterodimerises with MID1 via coiled-coil motifs and recruits the PP2A regulatory subunit Alpha4 through its B-boxes, with dimerisation required for the MID-Alpha4 complex to associate with microtubules (PMID:11806752). As a ligase it ubiquitinates astrin on Lys409 to drive its proteasomal degradation at intercellular bridge microtubules during cytokinesis, such that MID2 loss stabilizes astrin and produces cytokinetic failure, multinucleation, and cell death (PMID:26748699). MID2 further recruits LRRK2 to microtubules by binding a regulatory loop (residues 911-919) for ubiquitination and degradation, with LRRK2 Ser910/Ser935 phosphorylation governing the choice between cytosolic 14-3-3 and microtubule-bound MID2, and MID2 restraining Rab29-driven LRRK2 kinase activity in an E3-ligase-dependent manner (PMID:35266954). Beyond degradative ubiquitination, MID2 mediates K63-linked ubiquitination of HIF1α at Lys214 to promote its nuclear translocation and transcriptional activity (PMID:38769340), and modulates Wnt/β-catenin signaling and EMT upstream of PP2A (PMID:32953821). In development, the vertebrate ortholog stabilizes apicobasally polarized microtubules during neural tube closure together with Mig12 (PMID:20534674). A separate set of findings describes a *S. cerevisiae* protein named Mid2 — an O-mannosylated plasma-membrane sensor that activates the PKC/MAPK cell-wall-integrity pathway by stimulating Rom2-dependent GTP loading of Rho1 (PMID:10330137, PMID:11113201) — which is a distinct protein and does not bear on the vertebrate MID2 mechanism.

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 1999 Medium

    Established that vertebrate MID2 is a cytoskeletal protein, defining its subcellular platform and pointing to the B30.2 domain as the microtubule-targeting determinant.

    Evidence Direct subcellular imaging with domain-deletion analysis

    PMID:10644436

    Open questions at the time
    • Did not define a molecular activity or substrate
    • Single lab, localization only
  2. 2002 Medium

    Resolved how MID2 is assembled at the cytoskeleton, showing it dimerises with MID1 and recruits the PP2A subunit Alpha4, with dimerisation required for microtubule association.

    Evidence Yeast two-hybrid and domain-specific deletion mapping

    PMID:11806752

    Open questions at the time
    • Interactions not validated in a structural or reconstituted system
    • Functional consequence of Alpha4 recruitment not tested here
  3. 2010 Medium

    Connected MID2 to a developmental process, showing its loss disorganizes polarized microtubules during neural tube closure via cooperation with Mig12.

    Evidence Morpholino knockdown in Xenopus with live imaging and microtubule staining

    PMID:20534674

    Open questions at the time
    • Ubiquitin-ligase activity not implicated in this phenotype
    • Molecular mechanism of MT stabilization undefined
  4. 2013 Medium

    Defined the COS domain as required for cytoskeletal localization, linking a disease-associated missense mutation to mislocalization.

    Evidence GFP-tagged mutant expression and fluorescence microscopy in HEK293T

    PMID:24115387

    Open questions at the time
    • Single method (imaging) without biochemical validation
    • Effect on ligase activity not assessed
  5. 2015 High

    Identified the first MID2 ubiquitination substrate, establishing it as a RING E3 that degrades astrin at K409 to permit cytokinesis.

    Evidence Co-purification/MS, in vivo ubiquitination assay, K409A mutagenesis, and RNAi with cytokinesis phenotype

    PMID:26748699

    Open questions at the time
    • Does not establish ubiquitin chain linkage type on astrin
    • E2 partner not identified
  6. 2015 Medium

    Showed MID2 is functionally interchangeable with MID1 in a distinct cellular process, controlling lytic granule exocytosis in cytotoxic T cells.

    Evidence MID2 rescue in MID1-/- CTL, siRNA knockdown, exocytosis assay

    PMID:25924778

    Open questions at the time
    • Substrate or molecular mechanism in exocytosis unknown
    • Whether ligase activity is required not tested
  7. 2020 Medium

    Placed MID2 upstream of PP2A in Wnt/β-catenin signaling and EMT, linking it to phospho-regulation of PP2Ac and β-catenin stability.

    Evidence Overexpression/knockdown with TCF/LEF reporter, western blot, okadaic acid rescue, migration assay

    PMID:32953821

    Open questions at the time
    • Direct ubiquitination substrate in this pathway not identified
    • Mechanism linking MID2 to PP2Ac phosphorylation undefined
  8. 2020 Medium

    Identified an upstream transcriptional input, showing MORC4/STAT3 activates the MID2 promoter.

    Evidence Co-IP, ChIP-qPCR, dual-luciferase reporter

    PMID:32764967

    Open questions at the time
    • Physiological context of MID2 induction not established
    • Single lab
  9. 2022 High

    Defined a second physiological substrate and a regulatory switch, showing MID2 recruits LRRK2 (residues 911-919) to microtubules for degradation and restrains its kinase activity, with phospho-Ser910/935 partitioning LRRK2 between 14-3-3 and MID2.

    Evidence Quantitative MS interactome, co-IP, domain mapping, mutagenesis, proteasome inhibition, neurite outgrowth rescue

    PMID:35266954

    Open questions at the time
    • Ubiquitin chain type on LRRK2 not fully defined
    • In vivo relevance to LRRK2-associated disease not tested
  10. 2024 Medium

    Extended MID2 activity to non-degradative signaling, showing K63-linked ubiquitination of HIF1α at K214 drives its nuclear translocation.

    Evidence Co-IP, ubiquitination assay, K214 mutagenesis, nuclear fractionation

    PMID:38769340

    Open questions at the time
    • Single lab
    • Structural basis of K214 selection and chain assembly undefined

Open questions

Synthesis pass · forward-looking unresolved questions
  • It remains unknown how MID2 substrate selection, chain-linkage choice (K48- vs K63-linked), and its PP2A/Alpha4-dependent functions are integrated into a unified regulatory logic at the microtubule cytoskeleton.
  • No structural model of substrate engagement
  • Determinants of degradative vs non-degradative ubiquitination unresolved
  • Relationship between ligase activity and PP2A-dependent signaling not mechanistically connected

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0008092 cytoskeletal protein binding 3 GO:0016874 ligase activity 3 GO:0140096 catalytic activity, acting on a protein 3
Localization
GO:0005856 cytoskeleton 3 GO:0005829 cytosol 1
Pathway
R-HSA-162582 Signal Transduction 3 R-HSA-392499 Metabolism of proteins 3 R-HSA-1640170 Cell Cycle 1
Partners
ALPHA4ASTRINHIF1ALRRK2MID1

Evidence

Reading pass · 14 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2002 MID2 homo- and heterodimerises with MID1 via their coiled-coil motifs, and both proteins interact with Alpha4 (a regulatory subunit of PP2-type phosphatases) through their B-box domains; dimerisation is required for association of the MID-Alpha4 complex with microtubules. Yeast two-hybrid screens, domain-specific deletion analysis BMC cell biology Medium 11806752
1999 MID2 (FXY2) protein associates with microtubules in a manner dependent on its carboxy-terminal B30.2 domain. Subcellular localization by direct imaging; domain deletion analysis Genomics Medium 10644436
1999 MID2 is required for cell integrity signaling in response to pheromone in S. cerevisiae; it acts as a cell surface sensor upstream of the PKC/MAPK pathway and is highly O-mannosylated on its extracellular domain. Genetic epistasis (null mutant phenotypic analysis), dosage suppressor screen, immunofluorescence localization, biochemical O-mannosylation analysis Molecular and cellular biology High 10330137
2001 The C-terminal cytoplasmic domain of Mid2 interacts with Rom2 (a GEF for Rho1) in S. cerevisiae; mid2 mutant extracts are deficient in catalyzing GTP loading of Rho1 in vitro, indicating Mid2 functions to stimulate nucleotide exchange on Rho1 via Rom2. Additionally, O-mannosylation by Pmt2 is required for Mid2 signaling. Yeast two-hybrid, in vitro GTP-loading assay on Rho1, genetic analysis of pmt2 mutant Molecular and cellular biology High 11113201
2013 A missense mutation (p.Arg347Gln) in MID2 abolishes function of the COS domain, causing the GFP-tagged mutant protein to accumulate in the cytoplasm rather than binding the cytoskeleton, demonstrating that the COS domain is required for cytoskeletal localization of MID2. Transient expression of GFP-tagged mutant in HEK293T cells, fluorescence microscopy Human mutation Medium 24115387
2015 MID2 ubiquitinates astrin on lysine 409, targeting it for degradation during cytokinesis; MID2 and astrin co-localize at intercellular bridge microtubules. MID2 depletion stabilizes astrin, causes cytokinetic defects, multinucleation, and cell death. Expression of K409A mutant astrin phenocopies MID2 depletion. Co-purification/MS, co-localization by immunofluorescence, in vivo ubiquitination assay, RNAi knockdown with phenotypic readout, site-directed mutagenesis (K409A) Cell reports High 26748699
2010 MID2 (Xenopus ortholog) is required for neural tube closure; its depletion by morpholino knockdown destabilizes and disorganizes apicobasally polarized microtubules in the neural plate. MID2 cooperates with its interacting protein Mig12 for microtubule stabilization during neural plate remodeling. Morpholino-mediated knockdown in Xenopus, live imaging, microtubule staining Development (Cambridge, England) Medium 20534674
2022 TRIM1/MID2 (the mammalian protein) acts as an E3 ubiquitin ligase that recruits LRRK2 to the microtubule cytoskeleton for ubiquitination and proteasomal degradation; TRIM1 binds LRRK2 at residues 911-919 within a regulatory loop. Phosphorylation of LRRK2 Ser910/Ser935 determines whether LRRK2 associates with cytoplasmic 14-3-3 or microtubule-bound TRIM1. TRIM1 prevents Rab29-mediated upregulation of LRRK2 kinase activity in an E3-ligase-dependent manner. Quantitative mass spectrometry-based interactome, co-IP, domain mapping, site-directed mutagenesis, proteasome inhibitor experiments, neurite outgrowth rescue assay The Journal of cell biology High 35266954
2020 MID2 overexpression increases phosphorylation of PP2Ac (without changing total PP2Ac levels), downregulates β-catenin and Wnt/β-catenin signaling, and inhibits EMT and cell migration; MID2 knockdown has opposite effects. PP2A inhibition by okadaic acid partially rescues β-catenin levels after MID2 knockdown, placing MID2 upstream of PP2A in Wnt regulation. Western blot, TCF/LEF luciferase reporter assay, siRNA knockdown, wound healing assay, immunofluorescence Annals of translational medicine Medium 32953821
2015 MID2 can functionally substitute for MID1 in controlling exocytosis of lytic granules in cytotoxic T cells; transfection of MID2 into MID1-/- CTL completely rescues granule exocytosis, and knockdown of MID2 inhibits exocytosis in both wild-type and MID1-/- CTL. MID2 transfection rescue in MID1-/- CTL, siRNA knockdown, exocytosis assay APMIS Medium 25924778
2024 TRIM1/MID2 binds HIF1α and mediates K63-linked ubiquitination at Lys214 (in the loop between PAS domains), which is required for HIF1α nuclear translocation and transcriptional activation; mutation of Lys214 abolishes these effects. Co-IP, ubiquitination assay, site-directed mutagenesis (K214 mutant), nuclear fractionation Oncogenesis Medium 38769340
2020 MORC4 interacts with STAT3 (confirmed by Co-IP) and promotes transcriptional activation of MID2 via two STAT3-binding sites in the MID2 promoter; MORC4 inhibition reduces STAT3 enrichment at the MID2 promoter. Co-IP (MORC4-STAT3 interaction), ChIP-qPCR, dual-luciferase reporter assay OncoTargets and therapy Medium 32764967
1999 MID2 protein is uniformly distributed through the plasma membrane and is highly O-mannosylated on its extracellular domain in S. cerevisiae. Immunofluorescence microscopy, biochemical glycosylation analysis Molecular and cellular biology Medium 10330137
2018 ER targeting of MID2 mRNA in yeast requires the RNA-binding protein Khd1p and is independent of the signal sequence of the encoded protein. Liposome-binding assay, ER fractionation, KHD1 deletion mutant analysis FEBS letters Medium 29772604

Source papers

Stage 0 corpus · 30 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2001 Wsc1 and Mid2 are cell surface sensors for cell wall integrity signaling that act through Rom2, a guanine nucleotide exchange factor for Rho1. Molecular and cellular biology 258 11113201
1999 Mid2 is a putative sensor for cell integrity signaling in Saccharomyces cerevisiae. Molecular and cellular biology 174 10330137
2002 MID1 and MID2 homo- and heterodimerise to tether the rapamycin-sensitive PP2A regulatory subunit, alpha 4, to microtubules: implications for the clinical variability of X-linked Opitz GBBB syndrome and other developmental disorders. BMC cell biology 114 11806752
2010 MID1 and MID2 are required for Xenopus neural tube closure through the regulation of microtubule organization. Development (Cambridge, England) 66 20534674
1999 MID2, a homologue of the Opitz syndrome gene MID1: similarities in subcellular localization and differences in expression during development. Human molecular genetics 56 10400986
1994 The MID2 gene encodes a putative integral membrane protein with a Ca(2+)-binding domain and shows mating pheromone-stimulated expression in Saccharomyces cerevisiae. Gene 47 7828875
2007 Functional analyses of the extra- and intracellular domains of the yeast cell wall integrity sensors Mid2 and Wsc1. FEBS letters 46 17761172
2024 CircTRIM1 encodes TRIM1-269aa to promote chemoresistance and metastasis of TNBC via enhancing CaM-dependent MARCKS translocation and PI3K/AKT/mTOR activation. Molecular cancer 39 38755678
2013 Targeted deep resequencing identifies MID2 mutation for X-linked intellectual disability with varied disease severity in a large kindred from India. Human mutation 34 24115387
2010 Characterization of sensor-specific stress response by transcriptional profiling of wsc1 and mid2 deletion strains and chimeric sensors in Saccharomyces cerevisiae. Omics : a journal of integrative biology 34 20958245
2015 The X-Linked-Intellectual-Disability-Associated Ubiquitin Ligase Mid2 Interacts with Astrin and Regulates Astrin Levels to Promote Cell Division. Cell reports 33 26748699
2014 Yeast endocytic adaptor AP-2 binds the stress sensor Mid2 and functions in polarized cell responses. Traffic (Copenhagen, Denmark) 32 24460703
1999 FXY2/MID2, a gene related to the X-linked Opitz syndrome gene FXY/MID1, maps to Xq22 and encodes a FNIII domain-containing protein that associates with microtubules. Genomics 30 10644436
2005 Ace2p contributes to fission yeast septin ring assembly by regulating mid2+ expression. Journal of cell science 29 16317047
2019 Emerging Roles of the TRIM E3 Ubiquitin Ligases MID1 and MID2 in Cytokinesis. Frontiers in physiology 26 30941058
2013 The cell wall sensors Mtl1, Wsc1, and Mid2 are required for stress-induced nuclear to cytoplasmic translocation of cyclin C and programmed cell death in yeast. Oxidative medicine and cellular longevity 24 24260614
2020 MORC4 Promotes Chemoresistance of Luminal A/B Breast Cancer via STAT3-Mediated MID2 Upregulation. OncoTargets and therapy 21 32764967
1999 RGD1 genetically interacts with MID2 and SLG1, encoding two putative sensors for cell integrity signalling in Saccharomyces cerevisiae. Yeast (Chichester, England) 18 10590461
2022 The E3 ligase TRIM1 ubiquitinates LRRK2 and controls its localization, degradation, and toxicity. The Journal of cell biology 17 35266954
2015 Mid1/Mid2 expression in craniofacial development and a literature review of X-linked opitz syndrome. Molecular genetics & genomic medicine 17 26788540
2022 Septin filament compaction into rings requires the anillin Mid2 and contractile ring constriction. Cell reports 15 35443188
1998 Overexpression of MID2 suppresses the profilin-deficient phenotype of yeast cells. Molecular microbiology 12 9720869
2018 Molecular characterization of Rhodeus uyekii tripartite motif protein 1 (TRIM1) involved in IFN-γ/LPS-induced NF-κB signaling. Fish & shellfish immunology 9 29747011
2024 An E3 ligase TRIM1 promotes colorectal cancer progression via K63-linked ubiquitination and activation of HIF1α. Oncogenesis 8 38769340
2020 MID1 and MID2 regulate cell migration and epithelial-mesenchymal transition via modulating Wnt/β-catenin signaling. Annals of translational medicine 6 32953821
2018 Signal sequence-independent targeting of MID2 mRNA to the endoplasmic reticulum by the yeast RNA-binding protein Khd1p. FEBS letters 6 29772604
2015 MID2 can substitute for MID1 and control exocytosis of lytic granules in cytotoxic T cells. APMIS : acta pathologica, microbiologica, et immunologica Scandinavica 5 25924778
2024 Expanding the genetic and phenotypic spectrum of TRAPPC9 and MID2-related neurodevelopmental disabilities: report of two novel mutations, 3D-modelling, and molecular docking studies. Journal of human genetics 3 38467738
2020 A male infant with Xq22.2q22.3 duplication containing PLP1 and MID2. Molecular genetics & genomic medicine 2 31951325
2023 The Molecular and Function Characterization of Porcine MID2. Animals : an open access journal from MDPI 0 37760252

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