Affinage

RIPOR2

Rho family-interacting cell polarization regulator 2 · UniProt Q9Y4F9

Length
1068 aa
Mass
118.5 kDa
Annotated
2026-06-10
19 papers in source corpus 15 papers cited in narrative 15 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 8/9 claims corpus-supported (89%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

RIPOR2 (FAM65B) is a RhoA-regulatory cytoskeletal protein that controls cell polarity, migration, and differentiation across multiple cell types (PMID:23241886, PMID:25588844). It binds the small GTPase RhoA through a noncanonical domain and represses RhoA activity by reducing its GTP loading, thereby negatively regulating chemokine-induced T cell adhesion, polarization, and migration (PMID:23241886). This activity is regulated by phosphorylation: chemokine-induced phosphorylation lowers RIPOR2 affinity for RhoA and drives its relocation from the plasma membrane to the cytosol, with the degree of phosphorylation tuning RhoA-dependent actin polymerization and T cell migration in vivo (PMID:30254631). In neutrophils, RIPOR2 accumulates at the leading edge downstream of PLCβ/PI3Kγ signaling, is stabilized by phosphorylation-dependent 14-3-3 binding, and restrains RhoA and phosphorylated myosin light chain at the cell front to maintain chemotactic directionality (PMID:25588844). Its expression is transcriptionally controlled by FOXO1 (PMID:23241886) and by the RAS/ERK pathway, the latter promoting multinucleation in melanoma cells (PMID:42100747). In cochlear hair cells, RIPOR2 oligomers form a circumferential ring near the basal taper of stereocilia, binding RhoC (which regulates its oligomerization) and Myh9, and are required for taperin organization and mechanotransduction (PMID:27269051, PMID:30280293); correct stereociliary localization, dependent on a PX membrane-targeting domain, is functionally essential, and disrupting mutations cause hearing loss (PMID:24958875, PMID:32631815). Aminoglycosides bind RIPOR2 and trigger its translocation from stereocilia, whereupon it engages the autophagy protein GABARAP to dysregulate autophagy and PINK1/Parkin-dependent mitophagy, driving hair cell death (PMID:36113482). In myogenic cells RIPOR2 localizes to the cytoskeleton and filopodia and forms a transient HDAC6–dysferlin tricomplex required for myoblast differentiation and fusion (PMID:24687993, PMID:17150207). RIPOR2 mutation causes autosomal dominant adult-onset progressive hearing loss (PMID:32631815).

Mechanistic history

Synthesis pass · year-by-year structured walk · 14 steps
  1. 2006 Medium

    Established the first cellular role of the protein by linking it to the cytoskeleton and myogenesis, before its RhoA connection was known.

    Evidence Immunofluorescence, overexpression with serial deletion mutants mapping filopodia activity to aa 55–113, and siRNA knockdown in C2C12 myoblasts

    PMID:17150207

    Open questions at the time
    • Molecular mechanism linking the protein to actin protrusions not defined
    • No binding partners identified at this stage
  2. 2012 High

    Defined the core molecular function as a noncanonical RhoA inhibitor under FOXO1 transcriptional control, answering how it negatively regulates T cell migration.

    Evidence Pulldown and RhoA GTP-loading assays with knockdown/overexpression and chemotaxis/adhesion assays in T lymphocytes

    PMID:23241886

    Open questions at the time
    • Structural basis of the noncanonical RhoA-binding domain not resolved
    • How RhoA binding is dynamically regulated not yet addressed
  3. 2014 High

    Connected the protein to muscle differentiation mechanistically by identifying a transient HDAC6–dysferlin tricomplex and an in vivo myogenic requirement.

    Evidence Reciprocal pulldowns in differentiating human muscle, HDAC inhibitor treatment, and zebrafish knockdown with myoseptal histology

    PMID:24687993

    Open questions at the time
    • Function of the tricomplex during fusion not biochemically defined
    • Whether RhoA regulation participates in myogenesis unknown
  4. 2014 High

    Identified a PX membrane-targeting domain and tied its disruption to mislocalization and hair cell loss, opening the cochlear/deafness chapter.

    Evidence Mutant protein localization, zebrafish morpholino knockdown, and cosegregation in a consanguineous kindred

    PMID:24958875

    Open questions at the time
    • Lipid specificity of the PX domain not characterized
    • Stereociliary function of the protein not yet defined
  5. 2015 High

    Explained how the RhoA inhibitor is spatially controlled at the migrating cell front via phosphorylation-dependent 14-3-3 binding and front-signaling pathways.

    Evidence Live imaging, fractionation, phosphorylation and 14-3-3 Co-IP, and KO chemotaxis/adhesion-under-flow assays in neutrophils

    PMID:25588844

    Open questions at the time
    • Kinase responsible for leading-edge phosphorylation not identified
    • Quantitative coupling between RhoA suppression and pMLC localization unresolved
  6. 2016 High

    Resolved the supramolecular organization in stereocilia, identifying RhoC as an oligomerization regulator and linking the protein to mechanotransduction.

    Evidence STORM super-resolution imaging, yeast two-hybrid, oligomerization biochemistry, and KO mouse mechanotransduction recordings

    PMID:27269051

    Open questions at the time
    • Atomic structure of the ring oligomer not determined
    • Mechanism by which RhoC controls oligomerization not detailed
  7. 2016 Medium

    Showed expression level acts as a proliferation/mitotic switch, with a HDAC6–14-3-3 complex forming upon arrest.

    Evidence Overexpression in transformed cells with cell-cycle analysis, Co-IP, and forced expression/knockdown in primary T cells

    PMID:27556504

    Open questions at the time
    • Mechanism of spindle disruption not defined
    • Physiological relevance of mitotic block in non-overexpression contexts unclear
  8. 2018 High

    Demonstrated phosphorylation as the master switch coupling chemokine signaling to RhoA release and membrane-to-cytosol shuttling in vivo.

    Evidence Conditional KO mouse, phosphorylation and RhoA-affinity assays, fractionation, and intranodal two-photon microscopy

    PMID:30254631

    Open questions at the time
    • Identity of the chemokine-activated kinase not established
    • Phosphosite mapping not fully resolved in this context
  9. 2018 Medium

    Placed the protein in a cardiac autophagy circuit as a PINK1 phosphorylation substrate at S46.

    Evidence In vitro kinase assay, cardiac transgenic mouse models, and I/R injury with autophagy flux assays

    PMID:30349076

    Open questions at the time
    • Finding embedded in a circRNA study; direct cardiac function of the protein not independently confirmed
    • Whether S46 phosphorylation links to RhoA regulation unknown
  10. 2018 Medium

    Identified Myh9 as a stereociliary partner and linked the protein to kinocilium positioning and cytoskeletal protein stability.

    Evidence Co-IP, western blotting, and immunofluorescence in Ripor2-deficient mouse cochlea

    PMID:30280293

    Open questions at the time
    • Mechanism by which loss reduces Myh9 protein despite higher mRNA unresolved
    • Direct vs indirect effect on tubulin acetylation not separated
  11. 2020 Medium

    Established a human disease link, showing a dominant in-frame deletion causes hearing loss through mislocalization and loss of rescue function.

    Evidence Ex vivo mutant localization, rescue assay in deficient hair cells, and exome co-segregation in 12 families

    PMID:32631815

    Open questions at the time
    • Dominant-negative vs haploinsufficiency mechanism not distinguished
    • Why hearing loss is adult-onset and progressive not explained
  12. 2022 High

    Defined an ototoxicity mechanism: aminoglycoside binding triggers RIPOR2 translocation that hijacks GABARAP-dependent autophagy and PINK1/Parkin mitophagy to kill hair cells.

    Evidence Gentamicin-binding assay, live imaging, RIPOR2–GABARAP Co-IP, and RIPOR2/GABARAP/PINK1/Parkin KO/KD mice with ABR readouts

    PMID:36113482

    Open questions at the time
    • Structural site of gentamicin binding not mapped
    • How translocation switches RIPOR2 from cytoskeletal to autophagy function unknown
  13. 2025 Medium

    Dissected ototoxic pathways, showing aminoglycoside-induced RIPOR2 translocation is independent of phosphatidylserine externalization and distinct from cisplatin-induced death.

    Evidence Time-course live imaging and pharmacological/genetic dissection in wild-type hair cells

    PMID:40842562

    Open questions at the time
    • Molecular trigger separating the two death pathways not identified
    • Single-lab mechanistic dissection
  14. 2026 Medium

    Identified RAS/ERK as a transcriptional driver of RIPOR2 with a functional role in melanoma multinucleation.

    Evidence Single-nucleus RNA-seq, RAS/ERK activation in chicken embryo, and overexpression in human melanoma lines

    PMID:42100747

    Open questions at the time
    • Mechanism linking RIPOR2 to multinucleation not defined
    • Whether RhoA regulation underlies this phenotype untested

Open questions

Synthesis pass · forward-looking unresolved questions
  • How RIPOR2's distinct functional modes — cytoskeletal RhoA inhibition, stereociliary ring formation, and autophagy regulation — are mechanistically interconverted, and the structural basis of its noncanonical RhoA binding and oligomerization, remain open.
  • No atomic structure of RIPOR2 or its RhoA/RhoC complexes
  • Kinases governing context-specific phosphorylation incompletely defined
  • Unifying principle relating its membrane vs cytosol vs translocated states unresolved

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0008092 cytoskeletal protein binding 2 GO:0060090 molecular adaptor activity 2 GO:0098772 molecular function regulator activity 2
Localization
GO:0005886 plasma membrane 3 GO:0005829 cytosol 2 GO:0005856 cytoskeleton 2
Pathway
R-HSA-162582 Signal Transduction 3 R-HSA-1266738 Developmental Biology 2 R-HSA-9612973 Autophagy 1
Partners
DYSFGABARAPHDAC6MYH9PINK1RHOARHOCYWHAB
Complex memberships
RIPOR2–HDAC6–dysferlin tricomplex

Evidence

Reading pass · 15 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2012 FAM65B (RIPOR2) is a transcriptional target of FOXO1 that binds the small GTPase RhoA via a noncanonical domain and represses RhoA activity by decreasing its GTP loading, thereby negatively regulating chemokine-induced T cell adhesion, morphological polarization, and migration. Pulldown assays, RhoA activity (GTP-loading) assays, mRNA knockdown and overexpression in T lymphocytes, chemotaxis and adhesion assays Journal of immunology High 23241886
2014 FAM65B (RIPOR2) binds HDAC6 and dysferlin, forming a transient tricomplex (Fam65b–HDAC6–dysferlin) during myogenic cell differentiation; Fam65b expression is necessary for this complex to form, and its knockdown in developing zebrafish causes abnormal muscle with tears at the myosepta. Protein pulldowns in differentiating human muscle cells and regenerating muscle tissue, HDAC inhibitor treatment, zebrafish knockdown with histological readout FASEB journal High 24687993
2014 FAM65B (RIPOR2) contains a PX membrane localization domain required for plasma membrane targeting; a splice-site mutation causing deletion of 52 amino acids within this domain results in cytoplasmic inclusion body accumulation instead of membrane localization in stereocilia of cochlear hair cells, and knockdown in zebrafish reduces saccular hair cell and neuromast numbers. Mutant protein expression and localization (immunofluorescence), zebrafish fam65b morpholino knockdown, cosegregation in consanguineous kindred Proceedings of the National Academy of Sciences of the United States of America High 24958875
2015 FAM65B (RIPOR2) accumulates at the leading edge of neutrophils upon chemoattractant stimulation; phosphorylated FAM65B binds 14-3-3 family proteins, increasing its stability, and this accumulation depends on front-signal pathways (PLCβ and PI3Kγ). FAM65B deficiency in neutrophils increases RhoA activity and mislocalizes phosphorylated myosin light chain (pMLC) to the cell front, causing defects in chemotaxis directionality and adhesion to endothelial cells under flow. Live-cell imaging and subcellular fractionation in neutrophils, phosphorylation assays, Co-IP with 14-3-3 proteins, FAM65B-deficient cells, RhoA activity assays, chemotaxis and adhesion assays under flow Journal of cell science High 25588844
2016 Fam65b (RIPOR2) oligomers form a circumferential ring near the basal taper of stereocilia in murine cochlear hair cells as shown by STORM super-resolution microscopy; RhoC is a binding partner identified by yeast-two-hybrid and co-localizes with Fam65b in stereocilia; RhoC regulates Fam65b oligomerization; oligomerization and RhoC binding are required for Fam65b function. In Fam65b-deficient hair cells, taperin organization is disrupted and mechanotransduction is impaired. Stochastic optical reconstruction microscopy (STORM), biochemistry (oligomerization assays), yeast two-hybrid, co-localization, Fam65b-knockout mouse mechanotransduction recordings eLife High 27269051
2016 Forced FAM65B (RIPOR2) expression in transformed cells blocks mitosis by disrupting the mitotic spindle, causing G2 arrest and apoptosis; upon proliferation arrest, FAM65B forms a complex with HDAC6 and 14-3-3. In primary T cells, FAM65B is downregulated upon TCR engagement, and maintaining its expression blocks T cell proliferation. FAM65B overexpression in transformed cell lines, cell cycle analysis, Co-IP (FAM65B–HDAC6–14-3-3 complex), FAM65B knockdown and forced expression in primary T lymphocytes Oncotarget Medium 27556504
2018 Chemokine stimulation phosphorylates Fam65b (RIPOR2) in T lymphocytes; this phosphorylation decreases Fam65b affinity for RhoA and causes Fam65b to shuttle from the plasma membrane to the cytosol. Fam65b-deficient mice show increased spontaneous RhoA activation in resting T cells and defective intranodal T cell migration in vivo. The degree of Fam65b phosphorylation controls actin polymerization downstream of active RhoA and T cell migration in vitro. Conditional Fam65b-knockout mouse, phosphorylation assays, RhoA binding affinity measurements, subcellular fractionation, intranodal two-photon microscopy, in vitro migration assays Frontiers in immunology High 30254631
2018 In cardiomyocytes, PINK1 phosphorylates FAM65B (RIPOR2) at serine 46; phosphorylated FAM65B inhibits autophagy and cell death in the heart, placing FAM65B downstream of PINK1 in the ACR circRNA–PINK1–FAM65B axis regulating cardiac autophagy. In vitro kinase assay (PINK1 phosphorylating FAM65B at S46), cardiac-specific transgenic/overexpression mouse models, I/R injury model, autophagy flux assays Cell death and differentiation Medium 30349076
2018 Ripor2 (FAM65B) interacts with Myh9 (non-muscle myosin IIA, encoded by a known deafness gene) in cochlear hair cells; absence of Ripor2 reduces Myh9 protein abundance despite increased Myh9 mRNA, and causes aberrant kinocilium localization associated with reduced phosphorylated Myh9 and reduced acetylated alpha-tubulin in the kinocilium. Co-immunoprecipitation (Ripor2–Myh9 interaction), western blotting, immunofluorescence in Ripor2-deficient mouse cochlea Journal of molecular medicine Medium 30280293
2006 C6ORF32 (RIPOR2) localizes to the cellular cytoskeleton and filopodia in myogenic and non-myogenic cells; overexpression promotes formation of long membrane protrusions (filopodia), and a deletion analysis mapped filopodia-promoting activity to amino acids 55–113. Knockdown in C2C12 myoblasts causes a myogenic differentiation defect with decreased myogenin and myosin heavy chain expression. Immunofluorescence localization, overexpression with serial deletion mutants, siRNA knockdown in C2C12 cells with myogenic marker western blotting Developmental biology Medium 17150207
2022 Gentamicin binds to RIPOR2 and triggers its rapid translocation from stereocilia to the pericuticular area in murine hair cells; translocated RIPOR2 then interacts with the autophagy component GABARAP to disrupt autophagy. Reducing RIPOR2 or GABARAP expression completely prevents aminoglycoside-induced hair cell death and hearing loss in mice. Abolishing PINK1 or Parkin (mitophagy regulators) also prevents this hair cell death, placing RIPOR2-mediated autophagic dysfunction upstream of mitophagy-driven cell death. Drug-binding assay (gentamicin–RIPOR2), live-cell imaging of RIPOR2 translocation, Co-IP (RIPOR2–GABARAP interaction), RIPOR2/GABARAP/PINK1/Parkin knockout or knockdown mouse models, ABR hearing threshold measurements Developmental cell High 36113482
2020 An in-frame 12-nucleotide deletion in RIPOR2 causes autosomal dominant adult-onset progressive hearing loss; the mutant RIPOR2 protein shows aberrant localization in stereocilia of cochlear hair cells and fails to rescue morphological defects in RIPOR2-deficient hair cells, while wild-type RIPOR2 rescues them, establishing that correct stereociliary localization is functionally necessary. Ex vivo mutant protein expression and localization in hair cells, rescue assay in RIPOR2-deficient hair cells, exome sequencing co-segregation in 12 families Journal of medical genetics Medium 32631815
2025 Aminoglycoside-triggered RIPOR2 translocation and phosphatidylserine externalization in hair cells occur by independent mechanisms; cisplatin and aminoglycosides induce hair cell death via distinct molecular pathways (cisplatin does not replicate aminoglycoside-induced RIPOR2 translocation phenotype). Time-course live imaging of RIPOR2 translocation and PS externalization in wild-type hair cells with AG treatment, pharmacological and genetic dissection in hair cells Frontiers in cellular neuroscience Medium 40842562
2026 RIPOR2 is a positive transcriptional target of the RAS/ERK pathway in melanocyte precursors and human melanoma cells; ectopic RIPOR2 expression functionally promotes multinucleation in a chicken embryo in vivo model and in human melanoma-derived cell lines. Single-nucleus RNA sequencing, RAS/ERK pathway activation in chicken embryo model, RIPOR2 overexpression in human melanoma cell lines, multinucleation quantification iScience Medium 42100747
2025 RIPOR2 knockdown in T cells exacerbates exhaustion phenotypes (reduced IFN-γ secretion, increased PD-1); Ginsenoside F3 binds RIPOR2 (molecular docking confirmed), reverses exhaustion phenotypes, and synergizes with anti-PD-1 therapy in a mouse NSCLC model, identifying RIPOR2 as an immunometabolic regulator of T cell exhaustion. siRNA knockdown of RIPOR2 in in vitro T cell exhaustion model, proteomic profiling, molecular docking, in vivo NSCLC tumor model with anti-PD-1 co-treatment Phytomedicine Low 41418634

Source papers

Stage 0 corpus · 19 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2018 The circular RNA ACR attenuates myocardial ischemia/reperfusion injury by suppressing autophagy via modulation of the Pink1/ FAM65B pathway. Cell death and differentiation 221 30349076
2014 FAM65B is a membrane-associated protein of hair cell stereocilia required for hearing. Proceedings of the National Academy of Sciences of the United States of America 61 24958875
2012 Fam65b is a new transcriptional target of FOXO1 that regulates RhoA signaling for T lymphocyte migration. Journal of immunology (Baltimore, Md. : 1950) 42 23241886
2016 Murine Fam65b forms ring-like structures at the base of stereocilia critical for mechanosensory hair cell function. eLife 36 27269051
2006 C6ORF32 is upregulated during muscle cell differentiation and induces the formation of cellular filopodia. Developmental biology 33 17150207
2016 FAM65B controls the proliferation of transformed and primary T cells. Oncotarget 29 27556504
2014 Fam65b is important for formation of the HDAC6-dysferlin protein complex during myogenic cell differentiation. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 28 24687993
2018 Fam65b Phosphorylation Relieves Tonic RhoA Inhibition During T Cell Migration. Frontiers in immunology 25 30254631
2015 Front-signal-dependent accumulation of the RHOA inhibitor FAM65B at leading edges polarizes neutrophils. Journal of cell science 23 25588844
2022 RIPOR2-mediated autophagy dysfunction is critical for aminoglycoside-induced hearing loss. Developmental cell 19 36113482
2020 A RIPOR2 in-frame deletion is a frequent and highly penetrant cause of adult-onset hearing loss. Journal of medical genetics 17 32631815
2018 Ripor2 is involved in auditory hair cell stereociliary bundle structure and orientation. Journal of molecular medicine (Berlin, Germany) 13 30280293
2019 Faster regeneration associated to high expression of Fam65b and Hdac6 in dysferlin-deficient mouse. Journal of molecular histology 6 31218594
2025 Maternal Exposure to Environmentally Relevant Concentrations of Tris(2,4-di-tert-butylphenyl) Phosphate-Induced Developmental Toxicity in Zebrafish Offspring via Disrupting foxO1/ripor2 Signaling. Environmental science & technology 5 40087148
2019 Association of FAM65B, AGBL4, and CUX2 genetic polymorphisms with susceptibility to antituberculosis drug-induced hepatotoxicity: validation study in a Chinese Han population. Pharmacogenetics and genomics 5 30720667
2026 RIPOR2 promotes multinucleation of melanoma cells downstream of the RAS/ERK oncogenic pathway. iScience 0 42100747
2025 Aminoglycoside induces RIPOR2 translocation and phosphatidylserine externalization via distinct mechanisms. Frontiers in cellular neuroscience 0 40842562
2025 Ginsenoside F3 alleviates T cell exhaustion via RIPOR2-mediated immunometabolic reprogramming to potentiate anti-PD-1 therapy. Phytomedicine : international journal of phytotherapy and phytopharmacology 0 41418634
2023 RIPOR2: A new gene of non-syndromic cochleovestibular dysfunction, discrepancy between human pathology and animal models. Clinical genetics 0 37864412

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