Rho-related GTP-binding protein RhoD · UniProt O00212
RhoD is an atypical Rho-family GTPase that couples actin cytoskeletal remodeling to membrane trafficking, acting in many respects antagonistically to the canonical RhoA/RhoC contractile pathways (PMID:10229194, PMID:28486133). Constitutively active RhoD disassembles stress fibers and focal adhesions, suppresses cell migration, and arrests cytokinesis, opposing RhoA-driven contractility (PMID:10229194, PMID:28196728). It transduces these effects through a set of direct effectors: the formin splice variants hDia2C/mDia3C, which it recruits to drive actin polymerization at endosomes and at FGF-induced cytoneme-like protrusions (PMID:12577064, PMID:23034183), and the Arp2/3 nucleation-promoting factor WHAMM, which acts downstream of RhoD to control adhesion and migration (PMID:23087206). RhoD also recruits PAK6 to the plasma membrane to antagonize RhoC-ROCK-dependent contraction and blebbing, a circuit hijacked by the vaccinia virus protein F11 that inhibits RhoD (PMID:28486133), and it binds ZIP kinase in a GTP-dependent manner to suppress ZIPK-induced focal adhesion and FAK signaling (PMID:23454120). In membrane trafficking, RhoD localizes to early endosomes and the Golgi, regulating endosome motility via hDia2C and c-Src (PMID:12577064), PDGFβ-receptor internalization via the Rab5 effector Rabankyrin-5 (PMID:24102721), Golgi homeostasis and ER-to-plasma-membrane transport (PMID:25746724), and starvation-induced ATG9A trafficking from the trans-Golgi to phagophores during autophagy, in a complex with WHAMM (PMID:40143438). At plexin semaphorin receptors, RhoD binds the cytoplasmic Rho-GTPase-binding domain and destabilizes the effector-domain dimer, thereby antagonizing Rnd1-mediated plexin activation and Sema3A-induced axonal collapse (PMID:11784792, PMID:17916560). Biochemically, RhoD has elevated intrinsic GDP/GTP exchange activity, behaving as a constitutively active switch, and a unique N-terminal extension regulates its vesicle dynamics (PMID:29776664).
Establishing RhoD as a distinct Rho-family gene defined the molecular entity and its broad tissue distribution before any function was known.
Evidence cDNA library screening, sequencing, and Northern blot of human tissues
The first functional study showed RhoD opposes RhoA, answering whether this GTPase regulates the actin cytoskeleton and how it differs from canonical Rho proteins.
Evidence Constitutively active/dominant-negative mutants in fibroblasts and Xenopus embryos with migration and cytokinesis readouts
Identification of plexin-A1 binding established a non-cytoskeletal role in receptor regulation, showing RhoD antagonizes Rnd1 at semaphorin receptors.
Evidence GTPase interaction screens, dominant-active mutants, and axon collapse assays
RhoD was shown to re-route receptor signaling by inhibiting RhoA, framing it as a molecular switch between RhoA/ROCK and Galphaq-dependent pathways.
Evidence Pharmacological inhibitors, C3 exoenzyme, dominant mutants, and invasion assays
Linking RhoD to hDia2C and c-Src defined the first effector pathway, explaining how RhoD controls endosome motility along actin.
Evidence Co-IP/binding assays, overexpression, live imaging of endosome dynamics, dominant mutants
Mapping switch-II residues that dictate bacterial toxin specificity provided structure-function insight into RhoD's GTPase surface relative to RhoA.
Evidence Site-directed mutagenesis and in vitro toxin glucosylation/transglutamination assays
Two studies extended the endosome model: RhoD endosomes spatially target palmitoylated Src-family kinases, and structural work showed how RhoD destabilizes the plexin-B1 effector dimer.
Evidence siRNA knockdown with acylation-site mutants and colocalization; NMR plus 2.0 Å crystallography with binding assays
A burst of effector-mapping studies connected RhoD to WHAMM/FILIP1, mDia3C-driven cytoneme protrusions, and Diaph1-dependent G1/S progression and centrosome duplication.
Evidence Co-IP/pulldown, siRNA epistasis, FGF bead stimulation, live imaging, yeast two-hybrid, and mouse skin transgenics
PMID:22665057 PMID:23034183 PMID:23087206
RhoD was tied to Rab5-dependent PDGFβ-receptor internalization via Rabankyrin-5 and to GTP-dependent suppression of ZIPK contractile signaling, broadening its trafficking and anti-contractile roles.
Evidence Co-IP, colocalization, siRNA receptor internalization assays; yeast two-hybrid, co-IP, and FAK activity assays
Localization of RhoD to the Golgi and its requirement for ER-to-plasma-membrane transport extended its trafficking role to secretory organelle homeostasis.
Evidence Fluorescence localization, overexpression/knockdown, VSV-G trafficking assay
Two studies dissected RhoD's anti-contractile circuit, showing PAK6 recruitment antagonizes RhoC-ROCK (hijacked by vaccinia F11) and that RhoD globally restrains actin filament structures, migration, and proliferation.
Evidence Vaccinia infection, siRNA, co-IP, epistasis and contraction/blebbing assays; loss/gain-of-function with actin dynamics quantification across cell types
Structure-function dissection established RhoD as constitutively active with an N-terminal extension governing vesicle dynamics and distinct determinants of membrane versus vesicle localization.
Evidence Constitutively active and GDP-locked mutants with live imaging of vesicle fusion
Computational modeling proposed how RhoD inhibits and Rnd1 activates plexin via differential membrane interaction and an allosteric network, offering a mechanistic basis for opposing regulation.
Evidence Molecular dynamics simulations with structural analysis
RhoD was placed in autophagy, showing it drives ATG9A export from the Golgi to phagophores during starvation in a WHAMM- and domain-dependent manner.
Evidence Co-IP, BiFC, PUP-IT proximity labeling, RHOD knockout, domain-deletion mutants, autophagy flux assays
| Year | Finding | Method | Journal | Conf | PMIDs |
|---|---|---|---|---|---|
| 2003 | RhoD regulates early endosome motility by binding a novel splice variant of human Diaphanous, hDia2C, which is recruited onto early endosomes. RhoD and hDia2C together induce alignment of early endosomes along actin filaments and reduce their motility through membrane recruitment and activation of c-Src kinase, defining a signal transduction pathway in which hDia2C and c-Src are sequentially activated by RhoD. | Co-IP/binding assays, overexpression, live cell imaging of endosome dynamics, dominant-active/dominant-negative mutants | Nature cell biology | High | 12577064 |
| 2002 | RhoD directly binds the cytoplasmic domain of Plexin-A1 (as does Rnd1), and antagonizes Rnd1-mediated activation of Plexin-A1, thereby blocking Sema3A-induced cytoskeletal collapse and repulsion of sympathetic axons. | Binding assays (pulldown/interaction screens with large panel of GTPases), dominant-active mutant overexpression, axon collapse assays | The Journal of neuroscience | High | 11784792 |
| 2007 | RhoD (along with Rac1 and Rnd1) directly binds the cytoplasmic Rho GTPase-binding domain (RBD) of plexin-B1 via beta-strands 3 and 4 and a short alpha-helical segment (not the CRIB motif), and binding of any one of the three GTPases destabilizes the plexin-B1 effector domain dimer, suggesting a mechanism for receptor regulation. | Solution NMR spectroscopy, 2.0 Å resolution X-ray crystallography, in vitro binding assays | The Journal of biological chemistry | High | 17916560 |
| 1999 | Constitutively active RhoD(G26V) causes disassembly of actin stress fibers and focal adhesions, suppresses cell migration, induces multinucleation, and arrests cytokinesis in Xenopus embryos without affecting nuclear division. RhoD effects on stress fibers are antagonistic to RhoA. | Microinjection, transfection of constitutively active (G26V) and dominant-negative (T31K) mutants, phagokinetic track assay, Xenopus embryo injection | Oncogene | High | 10229194 |
| 2001 | RhoD (rhoDG26V) alone alters vesicular dynamics in the endocytic/recycling circuit and inhibits endothelial cell motility, demonstrating a dual role in vesicular movement and cell motility. | Live cell imaging of vesicular dynamics, overexpression of constitutively active mutant, cell motility assays | European journal of cell biology | Medium | 11484930 |
| 2002 | Activated RhoD (G26V) functions as a molecular switch that, by inhibiting endogenous RhoA, re-routes PAR-1 receptor signaling from Galpha12/13-RhoA/ROCK to Galphaq-PLC-Ca2+/CaM-MLCK-dependent cellular invasion pathway. | Pharmacological inhibitors, dominant-active/dominant-negative mutants, C3 exoenzyme, biochemical signaling assays, invasion assays | FASEB journal | Medium | 11919159 |
| 2007 | Fyn (palmitoylated SFK) colocalizes with RhoD-positive endosomes, and siRNA-mediated knockdown of RhoD inhibits the peripheral membrane targeting of both Fyn and palmitoylated Src (S3C/S6C), demonstrating that RhoD endosomes are required for spatial activation of palmitoylated SFKs. | siRNA knockdown, site-directed mutagenesis of acylation sites, fluorescence colocalization, immunofluorescence | Journal of cell science | Medium | 17623777 |
| 2012 | RhoD binds the actin nucleation-promoting factor WHAMM (which binds Arp2/3 complex) and FILIP1 (which binds filamin A); WHAMM acts downstream of RhoD in regulating cytoskeletal dynamics, and siRNA depletion of RhoD or WHAMM increases cell attachment and decreases cell migration. | Co-IP/pulldown binding assays, siRNA knockdown, cell migration assays, cell adhesion assays | Molecular biology of the cell | High | 23087206 |
| 2012 | RhoD induces two types of cytoneme-like cellular protrusions in response to FGF2/4/8 stimulation; activated RhoD specifically binds mDia3C and facilitates actin polymerization together with mDia3C, which localizes to tips/stems of protrusions; knockdown of either RhoD or mDia3 blocks protrusion formation. | Constitutively active mutant expression, bead stimulation assays, siRNA knockdown, co-IP binding assays, live cell imaging, fluorescence localization | Molecular biology of the cell | High | 23034183 |
| 2012 | RhoD is involved in G1/S-phase cell cycle progression and centrosome duplication; Diaph1 was identified as a novel effector of RhoD mediating G1/S progression; the effects on cell cycle and centrosome duplication are independent of each other. | Transgenic mouse model (skin), gain/loss-of-function in vitro, centriole overduplication assay in aphidicolin-arrested U2OS cells, yeast two-hybrid screen for Diaph1 interaction | Oncogene | Medium | 22665057 |
| 2013 | RhoD binds the Rab5 effector Rabankyrin-5, colocalizes with it on Rab5-positive endosomes, and siRNA depletion of RhoD interferes with internalization of the PDGFβ receptor and downstream signaling activation. | Co-IP/pulldown, fluorescence colocalization, siRNA knockdown, receptor internalization assays, signaling assays | Traffic | Medium | 24102721 |
| 2013 | RhoD interacts with ZIP kinase (ZIPK) in a GTP-dependent manner; co-expression of active RhoD suppresses ZIPK-induced stress fiber bundling, membrane blebbing, and focal adhesion reorganization; RhoD suppresses ZIPK-dependent FAK activity. | Yeast two-hybrid screen, co-IP, overexpression, GTP-dependence assay, FAK activity assays | Biochemical and biophysical research communications | Medium | 23454120 |
| 2015 | RhoD localizes to the Golgi apparatus and regulates Golgi homeostasis; manipulation of RhoD levels or activity (as well as its binding partner WHAMM) derails Golgi stack localization and severely impairs ER-to-plasma membrane vesicle trafficking as measured by VSV-G transport. | Fluorescence localization, overexpression/knockdown, VSV-G trafficking assay | Experimental cell research | Medium | 25746724 |
| 2017 | RhoD recruits PAK6 to the plasma membrane and uses it to antagonize RhoC signaling; vaccinia virus protein F11 inhibits RhoD, thereby relieving RhoD-mediated suppression of PAK6, which allows RhoC-ROCK-dependent cell contraction and blebbing. | Vaccinia infection model, siRNA knockdown, overexpression, co-IP, cell contraction/blebbing assays, epistasis analysis | Developmental cell | High | 28486133 |
| 2017 | RhoD depletion leads to increased actin filament structures (cortical actin, stress fibers, edge ruffles) and decreased cell migration and proliferation; ectopic RhoD expression produces a less dynamic, intertwined actin filament network across multiple cell types. | siRNA knockdown, overexpression, live cell imaging, actin dynamics quantification, migration assays | Experimental cell research | Medium | 28196728 |
| 2018 | Active GTP-bound RhoD conformation is required for plasma membrane localization but not vesicle localization; intact GTPase activity is required for efficient fusion of RhoD-positive vesicles; RhoD has an elevated intrinsic GDP/GTP exchange activity (constitutively active); the unique N-terminal 14-amino-acid extension regulates vesicle dynamics, as its deletion causes vesicle clustering at the peripheral membrane. | Constitutively active and GDP-locked mutants, fluorescence localization, live cell imaging of vesicle fusion | European journal of cell biology | Medium | 29776664 |
| 2006 | A single amino acid in the switch II region determines substrate specificity of RhoD for bacterial toxins: phenylalanine 85 of RhoD (equivalent to serine 73 in RhoA) prevents glucosylation by Clostridium difficile toxin B and allows modification by C. sordellii lethal toxin; aspartate 76 of RhoD (equivalent to glutamate 64 in RhoA) determines specificity for transglutaminating toxins DNT/CNF1. | Site-directed mutagenesis, in vitro glucosylation/transglutamination assays with bacterial toxins | The Journal of biological chemistry | High | 16702216 |
| 2021 | Molecular dynamics simulations show that RhoD destabilizes the plexin dimerization interface (acting as a plexin inhibitor) while RND1 reinforces it (acting as an activator), due to differential interaction with the inner leaflet of the cell membrane; this difference is attributed to RhoD's short C-terminal tail and positively charged membrane interface, which alter an allosteric network involving the RBD, RBD linkers, and a buttress segment. | Molecular dynamics (MD) simulations with structural analysis | eLife | Low | 34114565 |
| 2025 | RHOD interacts with ATG9A (a transmembrane protein essential for autophagosome formation) and accompanies ATG9A trafficking from the Golgi toward phagophores upon starvation; starvation-induced RHOD elevation causes Golgi fragmentation to promote ATG9A vesicle export from the trans-Golgi network; WHAMM forms a complex with RHOD and participates in this process in a RHOD-dependent manner; RHOD mutants lacking the exon II-containing effector region (required for ATG9A binding) or the CAAX box (required for membrane targeting) fail to stimulate ATG9A trafficking and autophagosome formation. | Co-IP, bimolecular fluorescence complementation (BiFC), PUP-IT proximity labeling, RHOD knockout, domain-deletion mutants, autophagy flux assays | Autophagy | High | 40143438 |
| 1997 | A novel human cDNA (rhoHP1/RHOD) was isolated from a human placenta cDNA library encoding a 210 amino acid protein with 50-54% amino acid identity to Rho family members; Northern analysis detected ~1.2 kb transcript in heart, placenta, liver, skeletal muscle, pancreas and other tissues. | cDNA library screening, sequencing, Northern blot | Biochimica et biophysica acta | Low | 9116026 |
| Year | Title | Journal | Citations | PMID |
|---|---|---|---|---|
| 2003 | RhoD regulates endosome dynamics through Diaphanous-related Formin and Src tyrosine kinase. | Nature cell biology | 175 | 12577064 |
| 2002 | Antagonistic effects of Rnd1 and RhoD GTPases regulate receptor activity in Semaphorin 3A-induced cytoskeletal collapse. | The Journal of neuroscience : the official journal of the Society for Neuroscience | 139 | 11784792 |
| 2007 | Binding of Rac1, Rnd1, and RhoD to a novel Rho GTPase interaction motif destabilizes dimerization of the plexin-B1 effector domain. | The Journal of biological chemistry | 113 | 17916560 |
| 2000 | Acute onset hemoglobinemia and/or hemoglobinuria and sequelae following Rh(o)(D) immune globulin intravenous administration in immune thrombocytopenic purpura patients. | Blood | 109 | 10753830 |
| 1983 | Molecular characterization of the human red cell Rho(D) antigen. | The EMBO journal | 91 | 11894930 |
| 2007 | The membrane targeting and spatial activation of Src, Yes and Fyn is influenced by palmitoylation and distinct RhoB/RhoD endosome requirements. | Journal of cell science | 90 | 17623777 |
| 1998 | Fluorescence measurement of calcium transients in perfused rabbit heart using rhod 2. | The American journal of physiology | 72 | 9486280 |
| 1984 | Should chronic transfusions be matched for antigens other than ABO and Rho(D)? | Vox sanguinis | 63 | 6431696 |
| 1999 | Small GTPase RhoD suppresses cell migration and cytokinesis. | Oncogene | 53 | 10229194 |
| 2012 | RhoD regulates cytoskeletal dynamics via the actin nucleation-promoting factor WASp homologue associated with actin Golgi membranes and microtubules. | Molecular biology of the cell | 48 | 23087206 |
| 2002 | RhoA- and RhoD-dependent regulatory switch of Galpha subunit signaling by PAR-1 receptors in cellular invasion. | FASEB journal : official publication of the Federation of American Societies for Experimental Biology | 48 | 11919159 |
| 2000 | Mitochondrial free calcium levels (Rhod-2 fluorescence) and ultrastructural alterations in neuronally differentiated PC12 cells during ceramide-dependent cell death. | The Journal of comparative neurology | 41 | 10982470 |
| 2012 | RhoD activated by fibroblast growth factor induces cytoneme-like cellular protrusions through mDia3C. | Molecular biology of the cell | 36 | 23034183 |
| 1967 | Relationship between Rh-o(D) zygosity and red cell Rh-o(D) antigen content in family members. | The Journal of clinical investigation | 35 | 4961263 |
| 1986 | Protective effect of the membrane skeleton on the immunologic reactivity of the human red cell Rho(D) antigen. | Journal of immunology (Baltimore, Md. : 1950) | 30 | 3086450 |
| 2013 | RhoD binds the Rab5 effector Rabankyrin-5 and has a role in trafficking of the platelet-derived growth factor receptor. | Traffic (Copenhagen, Denmark) | 29 | 24102721 |
| 2017 | The atypical Rho GTPase RhoD is a regulator of actin cytoskeleton dynamics and directed cell migration. | Experimental cell research | 27 | 28196728 |
| 2017 | RhoD Inhibits RhoC-ROCK-Dependent Cell Contraction via PAK6. | Developmental cell | 27 | 28486133 |
| 2001 | Rhod-2 based measurements of intracellular calcium in the perfused mouse heart: cellular and subcellular localization and response to positive inotropy. | Journal of biomedical optics | 27 | 11178577 |
| 2001 | Dual function of rhoD in vesicular movement and cell motility. | European journal of cell biology | 27 | 11484930 |
| 2013 | Interaction of RhoD and ZIP kinase modulates actin filament assembly and focal adhesion dynamics. | Biochemical and biophysical research communications | 20 | 23454120 |
| 2012 | RhoD participates in the regulation of cell-cycle progression and centrosome duplication. | Oncogene | 19 | 22665057 |
| 2011 | Dynamics of mitochondrial [Ca(2+)] measured with the low-Ca(2+)-affinity dye rhod-5N. | Cell calcium | 19 | 22133611 |
| 2006 | Exchange of a single amino acid switches the substrate properties of RhoA and RhoD toward glucosylating and transglutaminating toxins. | The Journal of biological chemistry | 19 | 16702216 |
| 2015 | RhoD is a Golgi component with a role in anterograde protein transport from the ER to the plasma membrane. | Experimental cell research | 18 | 25746724 |
| 2004 | Protein kinase C and myocardial calcium handling during ischemia and reperfusion: lessons learned using Rhod-2 spectrofluorometry. | The Thoracic and cardiovascular surgeon | 16 | 15192771 |
| 2014 | Atypical Rho GTPases RhoD and Rif integrate cytoskeletal dynamics and membrane trafficking. | Biological chemistry | 14 | 24622787 |
| 2009 | Investigation of whether the acute hemolysis associated with Rh(o)(D) immune globulin intravenous (human) administration for treatment of immune thrombocytopenic purpura is consistent with the acute hemolytic transfusion reaction model. | Transfusion | 13 | 19220820 |
| 1987 | Immunoreactivity of the Rho(D) antigen in cytoskeleton-free vesicles. | Transfusion | 11 | 3101248 |
| 1960 | Rho(D) genotype and red cell Rho(D) antigen content. | Science (New York, N.Y.) | 9 | 14422083 |
| 2021 | A putative structural mechanism underlying the antithetic effect of homologous RND1 and RhoD GTPases in mammalian plexin regulation. | eLife | 8 | 34114565 |
| 2018 | RhoD localization and function is dependent on its GTP/GDP-bound state and unique N-terminal motif. | European journal of cell biology | 7 | 29776664 |
| 1997 | Isolation of a novel human cDNA (rhoHP1) homologous to rho genes. | Biochimica et biophysica acta | 7 | 9116026 |
| 1982 | Identification of Rho(D) antigen in polyacrylamide gels by an enzyme-linked immunoassay. | Molecular immunology | 7 | 6810099 |
| 1996 | Risk estimation of ABO and Rho(D) incompatibility in persons with mono- and polyphyletic surnames in Monterrey, Mexico. Comparison with other Mexican populations. | Archives of medical research | 6 | 8696072 |
| 1978 | Studies on the characterization of the Rho(D) antigen. | Biochimica et biophysica acta | 6 | 103579 |
| 2017 | Effects of laser polarization on responses of the fluorescent Ca2+ indicator X-Rhod-1 in neurons and myelin. | Neurophotonics | 5 | 28612034 |
| 2024 | Rho(D) immune globulin shortage and fetal Rh(D) screening with cell-free DNA. | Current opinion in obstetrics & gynecology | 3 | 39774459 |
| 2016 | Toward Novel Diagnostics for Primary Open-Angle Glaucoma? An Association Study of Polymorphic Variation in Ras Homolog Family Member (A, B, C, D) Genes RHOA, RHOB, RHOC, and RHOD. | Omics : a journal of integrative biology | 3 | 27195967 |
| 2015 | Partial genome sequence of Thioalkalivibrio thiocyanodenitrificans ARhD 1(T), a chemolithoautotrophic haloalkaliphilic sulfur-oxidizing bacterium capable of complete denitrification. | Standards in genomic sciences | 3 | 26512309 |
| 2013 | The RhoD to centrosomal duplication. | Small GTPases | 3 | 23422264 |
| 1994 | Loss and reappearance of Rho(D) antigen on the red blood cells of an individual with acute myelogenous leukemia. | Immunohematology | 3 | 15945816 |
| 2025 | RHOD mediates ATG9A trafficking to promote autophagosome formation during autophagy in cancer. | Autophagy | 2 | 40143438 |
| 2020 | A New Rho(d) Map to Diffuse Gastric Cancer. | Cancer discovery | 2 | 32024682 |
| 2026 | Rho GTPase RhoD interacts with the Rho GEF Bud3 and is involved in aflatoxin biosynthesis, cell wall integrity, and pathogenicity in Aspergillus flavus. | Microbiological research | 0 | 41780129 |
| 1997 | [Immunoglobulin G monoclonal human anti-rhesus Rho(D) to prevent rhesus-incompatibility]. | Klinicheskaia meditsina | 0 | 9411052 |
| 1984 | [Isolation and determination of the molecular mass of the RHoD antigen in human erythrocytes]. | Bilten za hematologiju i transfuziju | 0 | 6441566 |
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