Affinage

DIAPH2

Protein diaphanous homolog 2 · UniProt O60879

Length
1101 aa
Mass
125.6 kDa
Annotated
2026-06-09
25 papers in source corpus 15 papers cited in narrative 16 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

DIAPH2/Dia2 has two distinct, evidence-supported identities in the available corpus: in budding yeast it functions as the F-box substrate-recognition subunit of an SCF(Dia2) E3 ubiquitin ligase that governs DNA replication, while the human ortholog acts as a cytoskeletal regulator (PMID:16421250, PMID:30926831). In yeast, Dia2 assembles an SCF complex through its F-box domain and binds replication origins, with its N-terminal TPR domain tethering the ligase to the replisome progression complex via the components Mrc1 and Ctf4 to concentrate it at replication forks (PMID:19913425, PMID:16421250). So positioned, SCF(Dia2) ubiquitylates the Mcm7 subunit of the CMG helicase at the end of replication to drive replisome disassembly through the Cdc48 segregase, the LRR motif being required for regulation of fork progression (PMID:19910927, PMID:26255844). Dia2 supports stable fork passage through damaged DNA and fragile regions such as the rDNA replication fork barrier, restrains premature S-phase entry, and promotes checkpoint recovery by aiding Rad53 deactivation and Mrc1 degradation; it additionally degrades Cdc6 in G1 to control origin licensing (PMID:16751663, PMID:23172854, PMID:23129771). Dia2 is itself cell-cycle-regulated: the Hect E3 ligase Tom1 ubiquitylates and degrades it during G1 and G2/M via positively charged residues in its N-terminal degradation/NLS domain, while S-phase checkpoint activation stabilizes it (PMID:19858292, PMID:22933573). In a separate functional arena, human DIAPH2 localizes to spindle microtubules and promotes microtubule polymerization through a microtubule-binding region outside its FH2 actin-nucleating domain, and mouse Diaph2 is expressed in cochlear outer hair cell stereocilia where a RhoA-activatable missense variant segregating with X-linked hearing loss impairs its function (PMID:30926831, PMID:36689403).

Mechanistic history

Synthesis pass · year-by-year structured walk · 8 steps
  1. 2006 High

    Establishing that Dia2 acts as an SCF E3 ligase subunit at replication origins reframed it from an uncharacterized factor into a cell-cycle regulator of replication and genome stability.

    Evidence Genetic deletion, cell-cycle analysis, and ChIP showing SCF assembly, origin binding, premature S-phase entry, and F-box-dependent damage phenotypes in budding yeast

    PMID:16421250 PMID:16751663

    Open questions at the time
    • Direct ubiquitylation substrates not yet identified at this stage
    • Mechanism linking origin binding to fork stability undefined
  2. 2007 Medium

    Identifying Yra1 as a recruiter addressed how Dia2 is delivered to chromatin.

    Evidence Affinity purification, reciprocal Co-IP, and ChIP showing Yra1-dependent origin recruitment of Dia2 and co-binding with Pol-delta subunit Hys2

    PMID:17452447

    Open questions at the time
    • Single-lab interaction
    • Relationship between Yra1- and TPR/replisome-mediated recruitment unresolved
  3. 2009 High

    Domain mapping resolved how Dia2 is physically coupled to the replisome and which motifs carry out distinct functions, defining the architecture of fork-associated ubiquitylation.

    Evidence Reciprocal Co-IP, two-hybrid, in vitro and in vivo ubiquitination, and domain deletion in two concurrent yeast studies showing TPR-Mrc1/Ctf4 tethering and LRR-dependent fork regulation

    PMID:19910927 PMID:19913425

    Open questions at the time
    • The decisive in vivo substrate driving fork regulation not pinned down here
    • Functional consequence of Mrc1/Ctf4 ubiquitylation ambiguous
  4. 2010 Medium

    Showing Dia2 is itself a degradation target under checkpoint control established a feedback layer regulating ligase abundance across the cell cycle.

    Evidence Protein stability assays, checkpoint-mutant analysis, and domain deletion identifying an N-terminal degradation/NLS domain and checkpoint-dependent stabilization

    PMID:19858292

    Open questions at the time
    • The responsible E3 ligase not identified at this stage
    • Single lab
  5. 2012 High

    Identifying Tom1 as the E3 that degrades Dia2, and Dia2/Tom1 roles in Cdc6 turnover and checkpoint recovery, connected ligase regulation to G1 origin licensing and damage response.

    Evidence In vivo ubiquitination, Co-IP, chromatin fractionation, kinase assays, and genetic suppression across three yeast studies

    PMID:22933573 PMID:23129771 PMID:23172854

    Open questions at the time
    • Whether Mrc1 degradation in recovery is direct vs indirect not fully resolved
    • Tom1 substrate-recognition structural basis undefined
  6. 2015 High

    Reconstituting Mcm7 ubiquitylation defined the central terminal-replication function: SCF(Dia2) triggers CMG helicase disassembly via Cdc48.

    Evidence In vitro and in vivo ubiquitylation, synthetic lethality with cdc48 allele, and TPR-tethering domain analysis showing Mcm7 as the key substrate, while ruling out replisome-specific Mrc1/Ctf4 degradation

    PMID:26255844

    Open questions at the time
    • How Mcm7 is selected at replication termination unresolved
    • Conservation of this pathway to the metazoan ortholog untested
  7. 2019 Medium

    Characterizing human DIAPH2 in mitosis revealed an FH2-independent microtubule-regulating activity, distinct from the yeast ubiquitin-ligase role.

    Evidence shRNA knockdown, live-cell imaging, in vitro MT polymerization assay, and a delta-FH2 mutant in colorectal cancer cells

    PMID:30793164 PMID:30926831

    Open questions at the time
    • Molecular partners of the non-FH2 MT-binding region unknown
    • Relationship between MT activity and migratory phenotype not mechanistically linked
  8. 2023 Medium

    Demonstrating stereocilia expression and a RhoA-activatable disease variant tied human DIAPH2 to actin-based sensory hair-cell function and X-linked hearing loss.

    Evidence Mouse cochlear immunohistochemistry, in vitro RhoA-activation functional assay, and CRISPR knockout/knockin mice with ABR measurements

    PMID:36689403

    Open questions at the time
    • Downstream cytoskeletal effectors in hair cells not identified
    • Single lab

Open questions

Synthesis pass · forward-looking unresolved questions
  • Whether the yeast SCF/replication functions and the metazoan cytoskeletal/formin functions reflect divergent activities of one conserved protein or fundamentally distinct proteins remains unresolved in the corpus.
  • No experiment tests both activities in a single ortholog
  • Domain conservation between yeast Dia2 and human DIAPH2 not addressed in timeline

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016874 ligase activity 2 GO:0140096 catalytic activity, acting on a protein 2 GO:0008092 cytoskeletal protein binding 1
Localization
GO:0005634 nucleus 1 GO:0005856 cytoskeleton 1 GO:0005929 cilium 1
Pathway
R-HSA-1640170 Cell Cycle 2 R-HSA-392499 Metabolism of proteins 2 R-HSA-69306 DNA Replication 2
Partners
CDC48CDC6CTF4MCM7MRC1RHOATOM1YRA1
Complex memberships
SCF(Dia2)replisome progression complex (associated)

Evidence

Reading pass · 16 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2009 The TPR domain at the amino terminus of yeast Dia2 tethers SCF(Dia2) to the replisome progression complex (RPC) by interacting with the RPC components Mrc1 and Ctf4, increasing the local concentration of the E3 ligase at DNA replication forks. Co-immunoprecipitation, two-hybrid interaction, domain deletion analysis Current biology : CB High 19913425
2009 SCF(Dia2) ubiquitinates replisome components Mrc1 and Ctf4 both in vivo and in vitro, and the leucine-rich repeat (LRR) motif of Dia2 is required for regulation of replisome progression while the TPR motif mediates interaction with replisome components and Dia2 stability. In vitro ubiquitination assay, in vivo ubiquitination, domain deletion analysis, yeast two-hybrid The EMBO journal High 19910927
2009 Dia2 localizes to replication forks and regulates replication fork progression under hydroxyurea treatment, as shown by ChIP-on-chip analysis. ChIP-on-chip The EMBO journal Medium 19910927
2015 SCF(Dia2) drives ubiquitylation of the Mcm7 subunit of the CMG helicase at the end of replication, and tethering via the TPR domain to the replisome progression complex increases the efficiency of this ubiquitylation both in vitro and in vivo, promoting CMG disassembly through a pathway requiring the Cdc48 segregase. SCF(Dia2) does NOT mediate replisome-specific degradation of Mrc1 and Ctf4. In vitro ubiquitylation assay, in vivo ubiquitylation, genetic epistasis (synthetic lethality with cdc48 allele), domain deletion Current biology : CB High 26255844
2006 Dia2 forms an SCF E3 ubiquitin ligase complex (SCF(Dia2)) in budding yeast and binds replication origins; deletion of DIA2 causes premature S-phase entry and DNA damage accumulation in S and G2/M phases; these defects require the F-box domain. Genetic deletion, cell cycle analysis, chromatin immunoprecipitation Molecular biology of the cell Medium 16421250
2006 Dia2 is required for stable passage of replication forks through damaged DNA and natural fragile regions (especially the rDNA replication fork barrier); loss of Dia2 activates the Rad53 checkpoint kinase, elevates DNA repair foci, and increases gross chromosomal rearrangements and extrachromosomal rDNA circles. Genetic deletion, systematic synthetic lethal screen (~4400 deletion mutants), checkpoint kinase assay, GCR assay Genetics High 16751663
2007 Yra1 (an mRNA export protein) is a Dia2 interaction partner; Yra1 and Dia2 co-bind replication origins and co-immunoprecipitate with DNA polymerase delta subunit Hys2; Dia2 binding to replication origins is significantly reduced when Yra1 association is compromised, suggesting Yra1 recruits Dia2 to chromatin. Affinity purification, co-immunoprecipitation, ChIP Molecular and cellular biology Medium 17452447
2010 Dia2 is itself targeted for ubiquitin-mediated proteolysis; activation of the S-phase checkpoint inhibits Dia2 protein degradation; an N-terminal domain (also required for nuclear localization) is necessary for degradation; Dia2 degradation does not involve an autocatalytic F-box mechanism. Protein stability assays, checkpoint mutant analysis, domain deletion, NLS addition rescue Molecular and cellular biology Medium 19858292
2012 The Hect-domain E3 ubiquitin ligase Tom1 ubiquitinates and degrades Dia2 during G1 and G2/M phases of the cell cycle; Tom1 binding to Dia2 is enhanced in G1 and reduced in S phase (when Dia2 is stabilized); Tom1 recognizes specific positively charged residues in the Dia2 degradation/NLS domain; loss of these residues blocks Tom1-mediated turnover and causes a G1-to-S delay. In vivo ubiquitination assay, co-immunoprecipitation, cell cycle staging, domain mutagenesis Molecular biology of the cell High 22933573
2012 SCF(Dia2) is required for checkpoint recovery from MMS-induced DNA damage: Dia2 promotes deactivation of the Rad53 checkpoint kinase and contributes to Mrc1 degradation during S-phase checkpoint recovery; checkpoint-defective mrc1 alleles suppress the MMS sensitivity and checkpoint recovery defect of dia2Δ cells. Genetic suppressor screen, Rad53 kinase assay, protein stability assay, MMS sensitivity assay Genetics Medium 23172854
2012 Dia2 and Tom1 each independently bind the C-terminal region of Cdc6 and control Cdc6 ubiquitination and degradation during G1 phase, acting separately from SCF(Cdc4); loss of Dia2 leads to aberrant Cdc6 and Mcm4 chromatin association in G1. Co-immunoprecipitation, in vivo ubiquitination assay, chromatin fractionation, genetic deletion The Journal of biological chemistry Medium 23129771
2011 Dia2 is involved in controlling assembly of the RSC chromatin-remodelling complex; in the absence of Dia2, RSC-mediated transcription regulation is impaired with abnormalities in nucleosome positioning. Genetic interaction screen, transcription assays, nucleosome positioning assay PloS one Low 21701592
2019 Human DIAPH2 controls spindle microtubule (MT) dynamics during M-phase in colorectal cancer cells independent of Cdc42 activity; DIAPH2 localizes to spindle MTs in metaphase; full-length DIAPH2 mediates a ~10-fold increase in MT polymerization in vitro independent of its FH2-domain actin-nucleating activity; a region outside the FH2 domain constitutes a second MT-binding site with distinct effects on MT dynamics. Stable knockdown (shRNA), live-cell imaging, in vitro MT polymerization assay, domain deletion mutant (ΔFH2) Scientific reports Medium 30926831
2019 Heterozygous DIAPH2 inactivation (CRISPR/Cas9) in HEK-293T cells shifts cells from a proliferative to a migratory phenotype, consistent with increased metastatic potential. CRISPR/Cas9 gene editing, proliferation and migration assays Carcinogenesis Low 30793164
2023 Mouse Diaph2 is expressed in the actin-rich stereocilia of cochlear outer hair cells; a missense variant in DIAPH2 segregating with X-linked hearing loss causes functional impairment of the protein upon RhoA-dependent activation in vitro. Immunohistochemistry (mouse cochlea), in vitro RhoA-activation assay, CRISPR/Cas9 knock-out/knock-in mice with ABR measurements PloS one Medium 36689403
2021 Absence of Dia2 in S. cerevisiae prolongs both S- and G2/M-phases, activates the S-phase checkpoint, and causes Ctf4 (a Dia2 substrate) to remain bound to chromatin for an extended period during these phases. Flow cytometry cell cycle analysis, chromatin fractionation/immunoprecipitation in dia2Δ cells Molecules (Basel, Switzerland) Low 35011329

Source papers

Stage 0 corpus · 25 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2009 The amino-terminal TPR domain of Dia2 tethers SCF(Dia2) to the replisome progression complex. Current biology : CB 65 19913425
2006 The F-box protein Dia2 overcomes replication impedance to promote genome stability in Saccharomyces cerevisiae. Genetics 52 16751663
2023 Long noncoding RNA DIAPH2-AS1 promotes neural invasion of gastric cancer via stabilizing NSUN2 to enhance the m5C modification of NTN1. Cell death & disease 46 37037818
2009 SCF(Dia2) regulates DNA replication forks during S-phase in budding yeast. The EMBO journal 45 19910927
2006 The F-box protein Dia2 regulates DNA replication. Molecular biology of the cell 41 16421250
2015 Tethering of SCF(Dia2) to the Replisome Promotes Efficient Ubiquitylation and Disassembly of the CMG Helicase. Current biology : CB 39 26255844
1977 An interpretation of human diaphorase isozymes in terms of three gene loci DIA1, DIA2 and DIA3. Annals of human genetics 34 563699
2012 The Hect domain E3 ligase Tom1 and the F-box protein Dia2 control Cdc6 degradation in G1 phase. The Journal of biological chemistry 20 23129771
2007 Yra1 is required for S phase entry and affects Dia2 binding to replication origins. Molecular and cellular biology 17 17452447
2019 DIAPH2 alterations increase cellular motility and may contribute to the metastatic potential of laryngeal squamous cell carcinoma. Carcinogenesis 15 30793164
2010 Activation of the S-phase checkpoint inhibits degradation of the F-box protein Dia2. Molecular and cellular biology 15 19858292
1998 Underexpression of a novel gene, dia2, impairs the transition of Dictyostelium cells from growth to differentiation. Biochemical and biophysical research communications 15 9813183
2012 Hect E3 ubiquitin ligase Tom1 controls Dia2 degradation during the cell cycle. Molecular biology of the cell 14 22933573
2012 The Saccharomyces cerevisiae F-box protein Dia2 is a mediator of S-phase checkpoint recovery from DNA damage. Genetics 14 23172854
2010 The intra-S phase checkpoint protein Tof1 collaborates with the helicase Rrm3 and the F-box protein Dia2 to maintain genome stability in Saccharomyces cerevisiae. The Journal of biological chemistry 13 21087929
2019 HOXD8/DIAPH2-AS1 epigenetically regulates PAX3 and impairs HTR-8/SVneo cell function under hypoxia. Bioscience reports 9 30626726
2019 The formin Drosophila homologue of Diaphanous2 (Diaph2) controls microtubule dynamics in colorectal cancer cells independent of its FH2-domain. Scientific reports 9 30926831
2021 DIAPH2, PTPRD and HIC1 Gene Polymorphisms and Laryngeal Cancer Risk. International journal of environmental research and public health 5 34299935
2011 Dia2 controls transcription by mediating assembly of the RSC complex. PloS one 5 21701592
2007 Involvements of a novel protein, DIA2, in cAMP signaling and spore differentiation during Dictyostelium development. Differentiation; research in biological diversity 5 17825085
2013 DIA-2, a polyherbal formulation ameliorates hyperglycemia and protein-oxidation without increasing the body weight in type II diabetic rats. European review for medical and pharmacological sciences 3 23426539
2023 In-depth genetic and molecular characterization of diaphanous related formin 2 (DIAPH2) and its role in the inner ear. PloS one 2 36689403
2015 Herbal formulation, DIA-2 and Rosiglitazone ameliorates hyperglycemia and hepatic steatosis in type 2 diabetic rats. European review for medical and pharmacological sciences 2 26367735
2021 The Effect of Dia2 Protein Deficiency on the Cell Cycle, Cell Size, and Recruitment of Ctf4 Protein in Saccharomyces cerevisiae. Molecules (Basel, Switzerland) 1 35011329
2024 DIAPH2 gene polymorphisms and laryngeal cancer risk in men. Journal of cancer research and clinical oncology 0 38844723

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