| 1994 |
RHAMM contains two HA-binding domains near the C-terminus defined by a B(X7)B motif (two basic amino acids flanking a seven-amino-acid stretch). Site-directed mutagenesis of K423, R431, and adjacent basic residues abolished HA binding; mutation of all B(X7)B motifs in recombinant RHAMM eliminated HA binding entirely. The same motif, when grafted onto the non-binding N-terminus of RHAMM, conferred HA binding, confirming this motif is necessary and sufficient for HA binding in RHAMM, CD44, and link protein. |
Site-directed mutagenesis of recombinant RHAMM fusion proteins, HA-Sepharose binding assays, chimeric protein construction, transblot analysis |
The EMBO journal |
High |
7508860
|
| 1994 |
RHAMM also contains a heparin-binding domain co-localizing with the HA-binding B(X7)B region at the C-terminus. Heparin at physiological concentrations (0.1 mg/ml) stimulated cell locomotion in a RHAMM-dependent manner (blocked by anti-RHAMM antibodies), while low concentrations (0.01 mg/ml) inhibited HA-induced locomotion independently of RHAMM. GST-RHAMM fusion protein bound biotin-labeled heparin in ligand blotting assays; deletion of HA-binding domains abolished heparin binding. |
Ligand blotting, HA-Sepharose/HP-Sepharose affinity chromatography, deletion analysis of fusion proteins, antibody neutralization of cell locomotion |
Journal of cellular biochemistry |
Medium |
7534313
|
| 1994 |
HA stimulation of ras-transformed fibroblast motility via RHAMM promotes rapid, transient protein tyrosine phosphorylation (within 1 min, dissipating by 10–15 min) and focal adhesion kinase (pp125FAK) phosphorylation/dephosphorylation, accompanied by rapid assembly and disassembly of focal adhesions (monitored by vinculin immunofluorescence). Tyrosine kinase inhibitors and microinjected anti-phosphotyrosine antibodies blocked both the phosphorylation and HA-stimulated motility. Anti-RHAMM antibodies reproduced the same tyrosine phosphorylation and focal adhesion turnover as HA. |
Immunoblot with anti-phosphotyrosine antibodies, immunofluorescence of vinculin, microinjection of antibodies, tyrosine kinase inhibitors (genistein, herbimycin A), time-course kinase assay |
The Journal of cell biology |
High |
7518470
|
| 1993 |
TGF-β1 stimulates cell locomotion by inducing transcription, synthesis, and membrane expression of RHAMM and coincident HA secretion. Anti-RHAMM antibodies and RHAMM HA-binding domain peptides suppressed TGF-β1-induced motility, placing RHAMM downstream of TGF-β1 in the motility pathway. |
Antibody neutralization, antisense oligonucleotides, exogenous TGF-β1 treatment, anti-RHAMM peptides, TGF-β1-transfected cell models |
The Journal of cell biology |
Medium |
7693717
|
| 1993 |
On ras-transformed fibroblasts, RHAMM (not CD44) is the primary mediator of HA-promoted cell locomotion. Anti-RHAMM antibodies completely inhibited HA-stimulated locomotion, whereas multiple anti-CD44 antibodies that block HA/CD44 interactions had no effect on locomotory responses to HA. |
Antibody neutralization assays, transblot HA-binding assay comparing CD44 and RHAMM, time-lapse locomotion assays |
Experimental cell research |
Medium |
7688314
|
| 1995 |
Overexpression of RHAMM transforms fibroblasts and causes spontaneous lung metastases. A dominant-suppressor RHAMM mutant reverses H-ras transformation and tumorigenicity; antisense reduction of RHAMM renders fibroblasts resistant to ras transformation. RHAMM acts downstream of ras, and loss of functional RHAMM ablates focal adhesion kinase (FAK) phosphorylation changes and prevents focal adhesion turnover in response to HA. |
Transfection overexpression, dominant-negative suppressor mutant, antisense transfection, in vivo tumor/metastasis assay, focal adhesion kinase phosphorylation analysis |
Cell |
High |
7541721
|
| 1996 |
Soluble RHAMM induces G2/M cell cycle arrest by suppressing Cdc2/Cyclin B1 expression through increased cdc2 mRNA degradation. Dominant-negative RHAMM mutants and antisense mRNA knockdown also decreased Cdc2 protein levels, linking RHAMM to mitotic entry control. |
Soluble RHAMM protein treatment, dominant-negative mutants, antisense knockdown, Western blot for Cdc2/Cyclin B1, mRNA stability assay, in vivo metastasis assay |
The Journal of experimental medicine |
Medium |
8666924
|
| 1998 |
RHAMM isoforms differ in subcellular localization: isoforms encoding exon 4 occur on the cell surface and in the cytoplasm; RHAMMv4 (encoding exon 4) is exclusively cytoplasmic. Intracellular RHAMMv4 regulates ERK1/2 activity: anti-RHAMM exon 4 antibodies block PDGF-induced ERK activation; a dominant-negative RHAMMv4 inhibits mutant active Ras-stimulated ERK activation and co-immunoprecipitates with MEK1 and ERK, indicating RHAMMv4 acts as an adaptor at the MEK–ERK level. Overexpression of RHAMMv4 constitutively activates ERK. |
Flow cytometry, confocal microscopy, epitope-tagging, co-immunoprecipitation with MEK1/ERK, dominant-negative expression, ERK kinase assay |
The Journal of biological chemistry |
High |
9556628
|
| 1999 |
Intracellular RHAMM/IHABP co-localizes with microtubules in interphase and dividing cells and is a novel microtubule-associated protein (MAP). Microtubule-binding region was mapped to the extreme N-terminus using deletion mutants fused to GFP; two subdomains are required for interphase microtubule binding, while one subdomain is sufficient for mitotic spindle binding. RHAMM/IHABP also interacts with actin filaments in vivo and in vitro. A calmodulin-binding site within residues 574–602 mediates Ca2+-dependent calmodulin binding. |
GFP fusion protein expression, microtubule co-sedimentation/binding assays, deletion mutant transfection, in vitro actin co-sedimentation, calmodulin-affinity chromatography |
Journal of cell science |
High |
10547355
|
| 2003 |
RHAMM localizes to centrosomes and is required for spindle pole stability. The N-terminus binds microtubules, while a separate C-terminal domain (72% identical to the dynein-interaction domain of Xklp2) is required for centrosomal targeting. Anti-RHAMM antibodies co-immunoprecipitate dynein intermediate chain (dynein IC) from Xenopus and HeLa extracts. Deregulation of RHAMM expression inhibits mitotic progression and disrupts spindle architecture. |
Immunofluorescence, co-immunoprecipitation of dynein IC, deletion mutant analysis, overexpression/knockdown cell biology, phylogenetic analysis |
Molecular biology of the cell |
High |
12808028
|
| 2006 |
Cell surface Rhamm is required for localization of CD44 to the cell surface, formation of CD44–ERK1,2 complexes, and nuclear targeting of activated ERK1,2 in fibroblasts. Rhamm−/− fibroblasts fail to resurface scratch wounds or invade HA-supplemented collagen gels; these defects are rescued by cell-surface-restricted recombinant Rhamm (bead-linked) or by expression of constitutively active MEK1, establishing that Rhamm acts upstream of ERK1,2 in this motogenic pathway. ERK1,2 activation and fibroblast migration/differentiation are also defective during skin wound repair in vivo in Rhamm−/− mice. |
Rhamm−/− genetic knockout mice, scratch wound assay, collagen gel invasion, co-immunoprecipitation (CD44–ERK1,2), subcellular fractionation, bead-linked recombinant Rhamm rescue, mutant active MEK1 rescue, in vivo excisional wound model |
The Journal of cell biology |
High |
17158951
|
| 2007 |
Cell surface Rhamm and CD44 form a complex with ERK1,2 that sustains high basal ERK1,2 activity and motility in invasive breast cancer cells, dependent on endogenous hyaluronan synthesis. CD44, Rhamm, and ERK1,2 uniquely co-immunoprecipitate and co-localize in invasive MDA-MB-231 and Ras-MCF10A cells but not in less invasive lines. Combined anti-CD44 + anti-Rhamm antibodies and MEK1 inhibitor showed less-than-additive effects, indicating action on a common pathway. |
Co-immunoprecipitation, co-localization by immunofluorescence, antibody neutralization, MEK1 inhibitor (PD098059), siRNA knockdown of HA synthase |
The Journal of biological chemistry |
High |
17392272
|
| 2007 |
RHAMM physically associates with the receptor tyrosine kinase RON (recepteur d'origine nantais) at the apex of ciliated airway epithelial cells. Oxidative degradation of apical HA produces low-molecular-weight HA fragments that signal via RHAMM and RON to increase ciliary beat frequency (CBF). A function-blocking anti-RHAMM antibody reduced the CBF response; co-immunoprecipitation confirmed physical RHAMM–RON association. |
Co-immunoprecipitation, immunohistochemistry, function-blocking antibodies, tyrosine kinase inhibitor (genistein), RON inhibitor (β-MSP), HA synthesis inhibition |
American journal of respiratory cell and molecular biology |
Medium |
17395888
|
| 2006 |
RHAMM, not CD44, mediates HA-induced vascular smooth muscle cell migration through a PI3K-dependent Rac activation pathway. siRNA knockdown of RHAMM inhibited both HA-induced migration and Rac activation; siRNA knockdown of CD44 inhibited RhoA activation without affecting migration. PI3K inhibitor LY294002 blocked HA-induced Rac activation and migration downstream of RHAMM. |
siRNA knockdown of RHAMM and CD44, Rho GTPase pull-down activity assays, PI3K inhibitor, Rho kinase inhibitor, blocking anti-CD44 antibody, migration assays |
Cardiovascular research |
Medium |
16934786
|
| 2008 |
Intracellular RHAMM associates with BRCA1 and BARD1; this association attenuates the mitotic-spindle-promoting activity of RHAMM. Extracellular RHAMM–CD44 partnering sustains CD44 surface display and enhances CD44-mediated ERK1/2 signaling. |
Co-immunoprecipitation (RHAMM–BRCA1, RHAMM–BARD1, RHAMM–CD44), cell surface display assays, ERK1/2 signaling assays (as described in cited primary studies reviewed here) |
Journal of cell science |
Low |
18354082
|
| 2010 |
Intracellular RHAMM(Δ163) regulates interphase and mitotic spindle microtubule stability through ERK1,2 activity. RHAMM−/− MEFs show strongly acetylated interphase microtubules, multipolar spindles, and aberrant cytokinesis, rescued by RHAMM or mutant active MEK1. RHAMM(Δ163) binds α- and β-tubulin via a C-terminal leucine zipper and directly binds ERK1 via a D-site motif (confirmed by co-IP and pulldown). RHAMM(Δ163)–ERK1/2–MEK1–tubulin complexes identified; RHAMM mutants defective in ERK1 binding fail to rescue microtubule defects. |
RHAMM−/− MEFs, mutant active MEK1 rescue, co-immunoprecipitation/pulldown of RHAMM–ERK1/2–MEK1–tubulin, in vitro tubulin polymerization assay, D-site mutant analysis, in vitro kinase assay |
The Journal of biological chemistry |
High |
20558733
|
| 2011 |
BRCA1 and RHAMM, together with AURKA and TPX2, cooperate in essential microtubule reorganization during MCF10A apicobasal polarization. BRCA1 facilitates this reorganization, while AURKA impairs it; RHAMM and TPX2 form a negative feedback loop regulating AURKA. Mechanistically, elevated RHAMM and TPX2 oppose AURKA activity at the mitotic cell cortex during polarization. |
MCF10A 3D polarization assay, siRNA knockdown of BRCA1/RHAMM/AURKA/TPX2, immunofluorescence, genetic association analysis in BRCA1 mutation carriers |
PLoS biology |
Medium |
22110403
|
| 2013 |
RHAMM expression is transcriptionally controlled by YAP/TEAD: YAP binds RHAMM promoter at specific sites and drives RHAMM transcription. Mevalonate pathway activity regulates YAP phosphorylation and nuclear-cytoplasmic localization, thereby controlling RHAMM transcription. Simvastatin inhibits breast cancer cell migration/invasion by reducing YAP-activated RHAMM transcription via geranylgeranylation, Rho GTPase activation, and actin cytoskeleton rearrangement, largely independently of MST/LATS kinase activity. |
Reporter assays (RHAMM promoter luciferase), ChIP for YAP/TEAD binding, siRNA knockdown of YAP, simvastatin treatment, geranylgeraniol rescue, YAP phosphorylation and localization analysis, in vitro/in vivo migration/invasion assays |
Proceedings of the National Academy of Sciences of the United States of America |
High |
24367099
|
| 2014 |
RHAMM is required for Aurora kinase A activation and correct TPX2 localization during mitotic spindle assembly. Silencing RHAMM delays spindle assembly kinetics, mislocalizes TPX2, and attenuates localized Aurora kinase A activation, reducing mitotic spindle length. The RHAMM–TPX2 complex requires a C-terminal basic leucine zipper in RHAMM and a domain including the nuclear localization signal in TPX2. |
siRNA silencing, live-cell imaging of spindle assembly kinetics, immunofluorescence of Aurora kinase A activity (pAURKA), co-immunoprecipitation of RHAMM–TPX2, deletion mutant analysis |
Cell cycle |
High |
24875404
|
| 2014 |
RHAMM acts as a co-activator of E2F1 to transcriptionally up-regulate fibronectin. E2F1 directly up-regulates RHAMM, which in turn enhances E2F1-mediated fibronectin expression and integrin-β1–FAK signaling, promoting cytoskeletal remodeling and tumor cell transmigration across endothelial layers. RHAMM depletion abolishes fibronectin expression and endothelial transmigration in E2F1-activated cells. |
Co-immunoprecipitation (E2F1–RHAMM), ChIP for E2F1 binding to RHAMM promoter, siRNA knockdown, fibronectin promoter reporter assay, transmigration assays, xenograft in vivo model |
The Journal of pathology |
Medium |
25042645
|
| 2015 |
In zebrafish heart regeneration, HA and Hmmr are required for epicardial epithelial-to-mesenchymal transition (EMT) and epicardial cell migration into the regenerating ventricle. Chemical inhibition of FAK or Src kinases (downstream effectors of Hmmr) prevented epicardial cell migration, implicating a HA/Hmmr/FAK/Src pathway in cardiac regeneration. |
Zebrafish morpholino knockdown of Hmmr, HA synthesis inhibitor, chemical inhibition of FAK and Src, ventricular resection model, proteomics identification of Hmmr |
Cardiovascular research |
Medium |
26156497
|
| 2016 |
RHAMM regulates planar germ cell division in the testis by associating with the mitotic spindle; loss of RHAMM from the spindle causes defective planar divisions of undifferentiated germ cells, leading to premature niche exit, germ cell depletion, hypofertility, and seminoma. RHAMM expression in testis is regulated by the testis-specific polyadenylation protein CFIm25, which is downregulated in human seminomas. |
Mouse models (Rhamm knockout, CFIm25 models), immunofluorescence of spindle-associated RHAMM, analysis of human seminoma specimens, CFIm25 expression studies |
Cancer research |
Medium |
27543603
|
| 2017 |
HMMR acts at centrosomes in a PLK1-dependent pathway that localizes active Ran and modulates cortical NuMA–dynein complex positioning to correct mispositioned mitotic spindles. Hmmr-knockout mice exhibit neonatal lethality with defective neural development and spindle orientation defects. HMMR overexpression induces phenotypes consistent with increased active Ran including spindle orientation defects. |
Hmmr-knockout mouse generation and phenotyping, immunofluorescence (active Ran, NuMA, dynein at cortex), PLK1 pathway epistasis, spindle orientation quantification in neural progenitors |
eLife |
High |
28994651
|
| 2018 |
HMMR acts as a nonmotor adaptor that dampens Eg5-mediated forces during mitosis by localizing TPX2 and promoting formation of inhibitory TPX2–Eg5 complexes. Genomic deletion or siRNA silencing of HMMR disturbs spindle microtubule organization, bipolar kinetochore attachments, and increases aneuploidy. A conserved HMMR motif with homology to kinesin Kif15 is required for interkinetochore tension and anaphase entry. HMMR defects are alleviated by chemical inhibition of Eg5 but not Kif15 silencing. |
siRNA silencing, HMMR genomic deletion, live-cell imaging, immunofluorescence (kinetochore tension, spindle architecture), chemical inhibition of Eg5 and Kif15 silencing, aneuploidy quantification, rescue with conserved motif constructs |
Molecular biology of the cell |
High |
29386294
|
| 2018 |
Phosphorylation of BACH1 during mitosis is required for its interaction with HMMR and CRM1 to stabilize mitotic spindle orientation. Mitosis-specific phosphorylations of BACH1 were identified by SILAC mass spectrometry; mutation of these phosphorylation sites abolished both spindle orientation rescue and HMMR interaction in BACH1-depleted cells. BACH1 loses chromatin/gene expression partners during mitosis but retains interaction with HMMR. |
SILAC mass spectrometry, co-immunoprecipitation (BACH1–HMMR), phosphomutant expression rescue, spindle orientation imaging, BACH1 knockdown and re-expression |
The Biochemical journal |
Medium |
29459360
|
| 2008 |
RHAMM expression is transcriptionally downregulated by p53. Reporter assays showed p53-dependent repression is mediated at the RHAMM promoter (including first exon and first intron). RHAMM protein levels peak in S phase and decrease before the G2/M mRNA peak, indicating post-transcriptional control in addition to transcriptional regulation during the cell cycle. |
Reporter assays (RHAMM promoter luciferase), p53-inducible transgenic cell system, nutlin-3/doxorubicin/paclitaxel treatment, RT-PCR and Western blot across cell cycle fractions |
Cell cycle |
Medium |
18971636
|
| 2001 |
In rat brain, RHAMM exists as multiple molecular weight forms (66, 75, 85–90 kDa) with differential subcellular distribution. The 75 kDa form is enriched in mitochondrial fractions and associates with mitochondrial membranes (retained in osmotically shocked mitochondria, liberated by alkali carbonate). Brain RHAMM binds calmodulin in a Ca2+-dependent manner via calmodulin-affinity chromatography. |
Subcellular fractionation, Western blotting, osmotic shock/alkali carbonate extraction of mitochondria, calmodulin-affinity chromatography, HA-Sepharose chromatography, double immunohistochemistry with cytochrome oxidase |
Journal of neuroscience research |
Medium |
11433424
|
| 2022 |
HMMR overexpression in mouse mammary epithelium increases Brca1-mutant tumorigenesis by activating AURKA, reducing ARPC2 localization at the mitotic cell cortex, promoting micronucleation, and activating cGAS-STING and non-canonical NF-κB signaling. Initial tumorigenic events include genomic instability, epithelial-to-mesenchymal transition, and tissue infiltration of tumor-associated macrophages. |
Transgenic mouse mammary epithelium overexpression of HMMR crossed with Brca1 mutant, immunofluorescence (AURKA, ARPC2 cortical localization), micronucleus assay, cGAS-STING pathway activation assays, NF-κB reporter/assays, macrophage infiltration histology |
Nature communications |
Medium |
35393420
|
| 2013 |
RHAMM interacts with ANKRD26 as identified by yeast two-hybrid and co-immunoprecipitation. Selective down-regulation of Hmmr in 3T3-L1 cells increased adipogenesis, indicating RHAMM suppresses this differentiation process. |
Yeast two-hybrid, co-immunoprecipitation, siRNA knockdown, adipogenesis assay (Oil Red O) |
PloS one |
Low |
22666460
|
| 2016 |
RHAMM forms an intracellular complex with β-catenin that protects β-catenin from degradation and supports its nuclear translocation, resulting in c-Myc activation and enhanced fibrosarcoma cell proliferation. LMWHA increases HT1080 cell growth in a RHAMM-dependent manner through this β-catenin/c-Myc axis. |
Co-immunoprecipitation (RHAMM–β-catenin), immunofluorescence, siRNA knockdown, transfection experiments with c-Myc reporter, cell proliferation assays |
Biochimica et biophysica acta |
Medium |
26825774
|
| 2020 |
RHAMM-dependent RHAMM/CD44 complexation is upregulated specifically by immobilized (end-on) HA but not by soluble HA in breast cancer cells. CD44/RHAMM co-localization and complexation were demonstrated by FRET microscopy and co-immunoprecipitation; this interaction is regulated in a cell-specific feedback loop via HA presentation format. |
FRET microscopy, co-immunoprecipitation, immunocytochemistry, comparison of soluble vs. immobilized HA substrates |
Acta biomaterialia |
Medium |
33091625
|
| 2023 |
HMMR promotes prostate cancer progression through a positive feedback loop: HMMR interacts with AURKA and stabilizes AURKA protein by inhibiting its ubiquitination-mediated degradation, which activates mTORC2/AKT signaling; activated AKT promotes E2F1-driven transcription of HMMR, forming a reinforcing loop. mTOR inhibitor partially antagonized HMMR-mediated tumor growth in vivo. |
Co-immunoprecipitation (HMMR–AURKA), ubiquitination assay, mTORC2/AKT phosphorylation analysis, E2F1 ChIP on HMMR promoter, gain/loss-of-function in vitro/in vivo, mTOR inhibitor treatment |
Cell death discovery |
Medium |
36750558
|
| 2023 |
HMMR forms a complex with FAK and SRC in the cytoplasm to activate NF-κB signaling, independently of membrane CD44 interaction, thereby sustaining CD47 ('don't eat me') signaling and enabling liver cancer immune evasion. HMMR knockout inhibited liver cancer growth and enhanced phagocytosis by macrophages; targeting HMMR enhanced anti-PD-1 treatment by recruiting CD8+ T cells. |
HMMR−/− liver cancer mouse model, co-immunoprecipitation (HMMR–FAK–SRC), NF-κB reporter assay, CD47 expression analysis, phagocytosis assays, anti-PD-1 combination in vivo |
Science advances |
Medium |
38838151
|
| 2020 |
In HCC progression driven by ER stress, HMMR is transcriptionally induced by the ER stress transcription factor CHOP and is ubiquitinated and degraded by the E3 ligase TRIM29. Dynamic TRIM29 expression during HCC progression regulates dynamic HMMR protein expression. HMMR alleviates ER stress by promoting autophagic lysosome activity. |
Co-immunoprecipitation, ubiquitination assay (TRIM29–HMMR), luciferase reporter assay (CHOP binding to HMMR promoter), ChIP, immunofluorescence (HMMR–autophagy markers), HBV-transgenic mouse model |
Cancer communications |
Medium |
37405956
|
| 2019 |
RHAMMB isoform (lacking the 15 aa encoded by alternative exon present in RHAMMA) but not RHAMMA promotes pancreatic tumor metastasis in vivo. RHAMMB upregulation in liver metastases correlates with higher EGFR expression, and EGFR knockdown abolished RHAMMB-driven metastasis, placing RHAMMB upstream of EGFR-dependent signaling. |
RNA-Seq isoform analysis of primary PNET and liver metastases, experimental metastasis mouse models, EGFR knockdown (siRNA), in vivo spontaneous metastasis assays |
Molecular cancer |
Medium |
31072393
|
| 2013 |
RHAMM cytoplasmic localization is required for maintenance of murine embryonic stem cell pluripotency via modulation of ERK1/2 and Aurora kinase A activity at microtubules. RHAMM was not detectable on the ESC cell surface. Hemizygous genomic deletion of Hmmr augmented differentiation and attenuated pluripotency; inhibition of ERK1/2 or Aurora kinase A rescued pluripotency in RHAMM+/− ESCs. |
Hemizygous Hmmr genomic deletion, immunofluorescence localization in ESCs, small-molecule kinase inhibitor screen, differentiation/pluripotency marker analysis |
PloS one |
Medium |
24019927
|