Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.


Logo for

J. Biol. Chem. 279 (26): 26991-27007

© 2004 by The American Society for Biochemistry and Molecular Biology, Inc.

CD44 Interaction with Na+-H+ Exchanger (NHE1) Creates Acidic Microenvironments Leading to Hyaluronidase-2 and Cathepsin B Activation and Breast Tumor Cell Invasion*

Lilly Y. W. Bourguignon{ddagger}§, Patrick A. Singleton{ddagger}, Falko Diedrich{ddagger}, Robert Stern¶, , and Eli Gilad{ddagger}

{ddagger}Department of Medicine, University of California, and Endocrine Unit, Veterans Affairs Medical Center and the Department of Pathology, University of California, San Francisco, California 94121

Abstract: We have explored CD44 (a hyaluronan (HA) receptor) interaction with a Na+-H+ exchanger (NHE1) and hyaluronidase-2 (Hyal-2) during HA-induced cellular signaling in human breast tumor cells (MDA-MB-231 cell line). Immunological analyses demonstrate that CD44s (standard form) and two signaling molecules (NHE1 and Hyal-2) are closely associated in a complex in MDA-MB-231 cells. These three proteins are also significantly enriched in cholesterol and ganglioside-containing lipid rafts, characterized as caveolin and flotillin-rich plasma membrane microdomains. The binding of HA to CD44 activates Na+-H+ exchange activity which, in turn, promotes intracellular acidification and creates an acidic extracellular matrix environment. This leads to Hyal-2-mediated HA catabolism, HA modification, and cysteine proteinase (cathepsin B) activation resulting in breast tumor cell invasion. In addition, we have observed the following: (i) HA/CD44-activated Rho kinase (ROK) mediates NHE1 phosphorylation and activity, and (ii) inhibition of ROK or NHE1 activity (by treating cells with a ROK inhibitor, Y27632, or NHE1 blocker, S-(N-ethyl-N-isopropyl) amiloride, respectively) blocks NHE1 phosphorylation/Na+-H+ exchange activity, reduces intracellular acidification, eliminates the acidic environment in the extracellular matrix, and suppresses breast tumor-specific behaviors (e.g. Hyal-2-mediated HA modification, cathepsin B activation, and tumor cell invasion). Finally, down-regulation of CD44 or Hyal-2 expression (by treating cells with CD44 or Hyal-2-specific small interfering RNAs) not only inhibits HA-mediated CD44 signaling (e.g. ROK-mediated Na+-H+ exchanger reaction and cellular pH changes) but also impairs oncogenic events (e.g. Hyal-2 activity, hyaluronan modification, cathepsin B activation, and tumor cell invasion). Taken together, our results suggest that CD44 interaction with a ROK-activated NHE1 (a Na+-H+ exchanger) in cholesterol/ganglioside-containing lipid rafts plays a pivotal role in promoting intracellular/extracellular acidification required for Hyal-2 and cysteine proteinase-mediated matrix degradation and breast cancer progression.

Received for publication October 29, 2003. Revision received April 15, 2004.

* This work was supported by United States Public Health Grants R01 CA66163, R01 CA 78633, and P01 AR39448 from the National Institutes of Health and a Veterans Affairs Merit Review grant. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ To whom correspondence and reprint requests should be addressed: Endocrine Unit (111N), Dept. of Medicine, University of California, San Francisco, and Veterans Affairs Medical Center, 4150 Clement St., San Francisco, CA 94121. Tel.: 415-221-4810 (Ext. 3321); Fax: 415-383-1638; E-mail: lillyb{at}

Pharmacological reversion of sphingomyelin-induced dendritic spine anomalies in a Niemann Pick disease type A mouse model.
A. I. Arroyo, P. G. Camoletto, L. Morando, M. Sassoe-Pognetto, M. Giustetto, P. P. Van Veldhoven, E. H. Schuchman, and M. D. Ledesma (2014)
EMBO Mol Med. 6, 398-413
   Abstract »    Full Text »    PDF »
NaV1.5 Na+ channels allosterically regulate the NHE-1 exchanger and promote the activity of breast cancer cell invadopodia.
L. Brisson, V. Driffort, L. Benoist, M. Poet, L. Counillon, E. Antelmi, R. Rubino, P. Besson, F. Labbal, S. Chevalier, et al. (2013)
J. Cell Sci. 126, 4835-4842
   Abstract »    Full Text »    PDF »
Membrane cholesterol modulates the hyaluronan-binding ability of CD44 in T lymphocytes and controls rolling under shear flow.
T. Murai, C. Sato, M. Sato, H. Nishiyama, M. Suga, K. Mio, and H. Kawashima (2013)
J. Cell Sci. 126, 3284-3294
   Abstract »    Full Text »    PDF »
KIAA1199, a deafness gene of unknown function, is a new hyaluronan binding protein involved in hyaluronan depolymerization.
H. Yoshida, A. Nagaoka, A. Kusaka-Kikushima, M. Tobiishi, K. Kawabata, T. Sayo, S. Sakai, Y. Sugiyama, H. Enomoto, Y. Okada, et al. (2013)
PNAS 110, 5612-5617
   Abstract »    Full Text »    PDF »
Regulation of the Na+/H+ Exchanger (NHE1) in Breast Cancer Metastasis.
S. R. Amith and L. Fliegel (2013)
Cancer Res. 73, 1259-1264
   Abstract »    Full Text »    PDF »
Murine Hyaluronidase 2 Deficiency Results in Extracellular Hyaluronan Accumulation and Severe Cardiopulmonary Dysfunction.
B. Chowdhury, R. Hemming, S. Hombach-Klonisch, B. Flamion, and B. Triggs-Raine (2013)
J. Biol. Chem. 288, 520-528
   Abstract »    Full Text »    PDF »
Role of Ion Channels and Transporters in Cell Migration.
A. Schwab, A. Fabian, P. J. Hanley, and C. Stock (2012)
Physiol Rev 92, 1865-1913
   Abstract »    Full Text »    PDF »
Renal interstitial hyaluronan: functional aspects during normal and pathological conditions.
S. Stridh, F. Palm, and P. Hansell (2012)
Am J Physiol Regulatory Integrative Comp Physiol 302, R1235-R1249
   Abstract »    Full Text »    PDF »
Intercellular Adhesion Molecule 1 Engagement Modulates Sphingomyelinase and Ceramide, Supporting Uptake of Drug Carriers by the Vascular Endothelium.
D. Serrano, T. Bhowmick, R. Chadha, C. Garnacho, and S. Muro (2012)
Arterioscler Thromb Vasc Biol 32, 1178-1185
   Abstract »    Full Text »    PDF »
Phosphoinositide Binding Differentially Regulates NHE1 Na+/H+ Exchanger-dependent Proximal Tubule Cell Survival.
B. G. Abu Jawdeh, S. Khan, I. Deschenes, M. Hoshi, M. Goel, J. T. Lock, K. Shinlapawittayatorn, G. Babcock, S. Lakhe-Reddy, G. DeCaro, et al. (2011)
J. Biol. Chem. 286, 42435-42445
   Abstract »    Full Text »    PDF »
Understanding the Dual Nature of CD44 in Breast Cancer Progression.
J. M. V. Louderbough and J. A. Schroeder (2011)
Mol. Cancer Res. 9, 1573-1586
   Abstract »    Full Text »    PDF »
Cortactin phosphorylation regulates cell invasion through a pH-dependent pathway.
M. A. O. Magalhaes, D. R. Larson, C. C. Mader, J. J. Bravo-Cordero, H. Gil-Henn, M. Oser, X. Chen, A. J. Koleske, and J. Condeelis (2011)
J. Cell Biol. 195, 903-920
   Abstract »    Full Text »    PDF »
Low Cholesterol Triggers Membrane Microdomain-dependent CD44 Shedding and Suppresses Tumor Cell Migration.
T. Murai, Y. Maruyama, K. Mio, H. Nishiyama, M. Suga, and C. Sato (2011)
J. Biol. Chem. 286, 1999-2007
   Abstract »    Full Text »    PDF »
V3 Versican Isoform Alters the Behavior of Human Melanoma Cells by Interfering with CD44/ErbB-dependent Signaling.
D. Hernandez, L. Miquel-Serra, M.-J. Docampo, A. Marco-Ramell, J. Cabrera, A. Fabra, and A. Bassols (2011)
J. Biol. Chem. 286, 1475-1485
   Abstract »    Full Text »    PDF »
Hyaluronan-CD44 Interaction Promotes c-Src-mediated Twist Signaling, MicroRNA-10b Expression, and RhoA/RhoC Up-regulation, Leading to Rho-kinase-associated Cytoskeleton Activation and Breast Tumor Cell Invasion.
L. Y. W. Bourguignon, G. Wong, C. Earle, K. Krueger, and C. C. Spevak (2010)
J. Biol. Chem. 285, 36721-36735
   Abstract »    Full Text »    PDF »
High-molecular-weight hyaluronan is a novel inhibitor of pulmonary vascular leakiness.
P. A. Singleton, T. Mirzapoiazova, Y. Guo, S. Sammani, N. Mambetsariev, F. E. Lennon, L. Moreno-Vinasco, and J. G. N. Garcia (2010)
Am J Physiol Lung Cell Mol Physiol 299, L639-L651
   Abstract »    Full Text »    PDF »
NHE1 promotes invadopodial ECM proteolysis through acidification of the peri-invadopodial space.
G. Busco, R. A. Cardone, M. R. Greco, A. Bellizzi, M. Colella, E. Antelmi, M. T. Mancini, M. E. Dell'Aquila, V. Casavola, A. Paradiso, et al. (2010)
FASEB J 24, 3903-3915
   Abstract »    Full Text »    PDF »
Reactive Oxygen Species and Hyaluronidase 2 Regulate Airway Epithelial Hyaluronan Fragmentation.
M. E. Monzon, N. Fregien, N. Schmid, N. S. Falcon, M. Campos, S. M. Casalino-Matsuda, and R. M. Forteza (2010)
J. Biol. Chem. 285, 26126-26134
   Abstract »    Full Text »    PDF »
HGF-induced invasion by prostate tumor cells requires anterograde lysosome trafficking and activity of Na+-H+ exchangers.
J. J. Steffan, B. C. Williams, T. Welbourne, and J. A. Cardelli (2010)
J. Cell Sci. 123, 1151-1159
   Abstract »    Full Text »    PDF »
Tumor Necrosis Factor-{alpha} Regulates Transforming Growth Factor-{beta}-dependent Epithelial-Mesenchymal Transition by Promoting Hyaluronan-CD44-Moesin Interaction.
E. Takahashi, O. Nagano, T. Ishimoto, T. Yae, Y. Suzuki, T. Shinoda, S. Nakamura, S. Niwa, S. Ikeda, H. Koga, et al. (2010)
J. Biol. Chem. 285, 4060-4073
   Abstract »    Full Text »    PDF »
Hyaluronan-CD44 Interactions in Cancer: Paradoxes and Possibilities.
B. P. Toole (2009)
Clin. Cancer Res. 15, 7462-7468
   Abstract »    Full Text »    PDF »
Inhibition of Functional Hyaluronan-CD44 Interactions in CD133-positive Primary Human Ovarian Carcinoma Cells by Small Hyaluronan Oligosaccharides.
M. G. Slomiany, L. Dai, L. B. Tolliver, G. D. Grass, Y. Zeng, and B. P. Toole (2009)
Clin. Cancer Res. 15, 7593-7601
   Abstract »    Full Text »    PDF »
Two Novel Functions of Hyaluronidase-2 (Hyal2) Are Formation of the Glycocalyx and Control of CD44-ERM Interactions.
C. Duterme, J. Mertens-Strijthagen, M. Tammi, and B. Flamion (2009)
J. Biol. Chem. 284, 33495-33508
   Abstract »    Full Text »    PDF »
Repression of NHE1 Expression by PPAR{gamma} Activation Is a Potential New Approach for Specific Inhibition of the Growth of Tumor Cells In vitro and In vivo.
A. P. Kumar, A. L. Quake, M. K. X. Chang, T. Zhou, K. S. Y. Lim, R. Singh, R. E. Hewitt, M. Salto-Tellez, S. Pervaiz, and M.-V. Clement (2009)
Cancer Res. 69, 8636-8644
   Abstract »    Full Text »    PDF »
Hyaluronan-CD44 Interaction with Protein Kinase C{epsilon} Promotes Oncogenic Signaling by the Stem Cell Marker Nanog and the Production of MicroRNA-21, Leading to Down-regulation of the Tumor Suppressor Protein PDCD4, Anti-apoptosis, and Chemotherapy Resistance in Breast Tumor Cells.
L. Y. W. Bourguignon, C. C. Spevak, G. Wong, W. Xia, and E. Gilad (2009)
J. Biol. Chem. 284, 26533-26546
   Abstract »    Full Text »    PDF »
Abrogating Drug Resistance in Malignant Peripheral Nerve Sheath Tumors by Disrupting Hyaluronan-CD44 Interactions with Small Hyaluronan Oligosaccharides.
M. G. Slomiany, L. Dai, P. A. Bomar, T. J. Knackstedt, D. A. Kranc, L. Tolliver, B. L. Maria, and B. P. Toole (2009)
Cancer Res. 69, 4992-4998
   Abstract »    Full Text »    PDF »
Proton Transport Inhibitors as Potentially Selective Anticancer Drugs.
Anticancer Res 29, 2127-2136
   Abstract »    Full Text »    PDF »
NLRP3/Cryopyrin Is Necessary for Interleukin-1{beta} (IL-1{beta}) Release in Response to Hyaluronan, an Endogenous Trigger of Inflammation in Response to Injury.
K. Yamasaki, J. Muto, K. R. Taylor, A. L. Cogen, D. Audish, J. Bertin, E. P. Grant, A. J. Coyle, A. Misaghi, H. M. Hoffman, et al. (2009)
J. Biol. Chem. 284, 12762-12771
   Abstract »    Full Text »    PDF »
Mislocalized Scaffolding by the Na-H Exchanger NHE1 Dominantly Inhibits Fibronectin Production and TGF-{beta} Activation.
A. Karydis, M. Jimenez-Vidal, S. P. Denker, and D. L. Barber (2009)
Mol. Biol. Cell 20, 2327-2336
   Abstract »    Full Text »    PDF »
Voltage-gated Sodium Channel Activity Promotes Cysteine Cathepsin-dependent Invasiveness and Colony Growth of Human Cancer Cells.
L. Gillet, S. Roger, P. Besson, F. Lecaille, J. Gore, P. Bougnoux, G. Lalmanach, and J.-Y. Le Guennec (2009)
J. Biol. Chem. 284, 8680-8691
   Abstract »    Full Text »    PDF »
Bicarbonate Increases Tumor pH and Inhibits Spontaneous Metastases.
I. F. Robey, B. K. Baggett, N. D. Kirkpatrick, D. J. Roe, J. Dosescu, B. F. Sloane, A. I. Hashim, D. L. Morse, N. Raghunand, R. A. Gatenby, et al. (2009)
Cancer Res. 69, 2260-2268
   Abstract »    Full Text »    PDF »
Interaction of monocarboxylate transporter 4 with {beta}1-integrin and its role in cell migration.
S. M. Gallagher, J. J. Castorino, and N. J. Philp (2009)
Am J Physiol Cell Physiol 296, C414-C421
   Abstract »    Full Text »    PDF »
Hyaluronan, CD44, and Emmprin Regulate Lactate Efflux and Membrane Localization of Monocarboxylate Transporters in Human Breast Carcinoma Cells.
M. G. Slomiany, G. D. Grass, A. D. Robertson, X. Y. Yang, B. L. Maria, C. Beeson, and B. P. Toole (2009)
Cancer Res. 69, 1293-1301
   Abstract »    Full Text »    PDF »
CD44 is of Functional Importance for Colorectal Cancer Stem Cells.
L. Du, H. Wang, L. He, J. Zhang, B. Ni, X. Wang, H. Jin, N. Cahuzac, M. Mehrpour, Y. Lu, et al. (2008)
Clin. Cancer Res. 14, 6751-6760
   Abstract »    Full Text »    PDF »
Chondroitin Sulfate E Fragments Enhance CD44 Cleavage and CD44-Dependent Motility in Tumor Cells.
K. N. Sugahara, T. Hirata, T. Tanaka, S. Ogino, M. Takeda, H. Terasawa, I. Shimada, J.-i. Tamura, G. B. ten Dam, T. H. van Kuppevelt, et al. (2008)
Cancer Res. 68, 7191-7199
   Abstract »    Full Text »    PDF »
Regulation of cell survival by Na+/H+ exchanger-1.
J. R. Schelling and B. G. Abu Jawdeh (2008)
Am J Physiol Renal Physiol 295, F625-F632
   Abstract »    Full Text »    PDF »
A mouse model of human mucopolysaccharidosis IX exhibits osteoarthritis.
D. C. Martin, V. Atmuri, R. J. Hemming, J. Farley, J. S. Mort, S. Byers, S. Hombach-Klonisch, R. Stern, and B. L. Triggs-Raine (2008)
Hum. Mol. Genet. 17, 1904-1915
   Abstract »    Full Text »    PDF »
Causes and Consequences of Increased Glucose Metabolism of Cancers.
R. J. Gillies, I. Robey, and R. A. Gatenby (2008)
J. Nucl. Med. 49, 24S-42S
   Abstract »    Full Text »    PDF »
Mouse Hyal3 encodes a 45- to 56-kDa glycoprotein whose overexpression increases hyaluronidase 1 activity in cultured cells.
R. Hemming, D. C. Martin, E. Slominski, J. I. Nagy, A. J. Halayko, S. Pind, and B. Triggs-Raine (2008)
Glycobiology 18, 280-289
   Abstract »    Full Text »    PDF »
JAK kinases promote invasiveness in VHL-mediated renal cell carcinoma by a suppressor of cytokine signaling-regulated, HIF-independent mechanism.
K. L. Wu, H. Miao, and S. Khan (2007)
Am J Physiol Renal Physiol 293, F1836-F1846
   Abstract »    Full Text »    PDF »
CD44 Regulates Hepatocyte Growth Factor-mediated Vascular Integrity: ROLE OF c-Met, Tiam1/Rac1, DYNAMIN 2, AND CORTACTIN.
P. A. Singleton, R. Salgia, L. Moreno-Vinasco, J. Moitra, S. Sammani, T. Mirzapoiazova, and J. G. N. Garcia (2007)
J. Biol. Chem. 282, 30643-30657
   Abstract »    Full Text »    PDF »
Inducible Hyaluronan Production Reveals Differential Effects on Prostate Tumor Cell Growth and Tumor Angiogenesis.
A. G. Bharadwaj, K. Rector, and M. A. Simpson (2007)
J. Biol. Chem. 282, 20561-20572
   Abstract »    Full Text »    PDF »
Heregulin-mediated ErbB2-ERK Signaling Activates Hyaluronan Synthases Leading to CD44-dependent Ovarian Tumor Cell Growth and Migration.
L. Y. W. Bourguignon, E. Gilad, and K. Peyrollier (2007)
J. Biol. Chem. 282, 19426-19441
   Abstract »    Full Text »    PDF »
Ability of Hyaluronidase 2 To Degrade Extracellular Hyaluronan Is Not Required for Its Function as a Receptor for Jaagsiekte Sheep Retrovirus.
V. Vigdorovich, A. D. Miller, and R. K. Strong (2007)
J. Virol. 81, 3124-3129
   Abstract »    Full Text »    PDF »
CD44-dependent Intracellular and Extracellular Catabolism of Hyaluronic Acid by Hyaluronidase-1 and -2.
H. Harada and M. Takahashi (2007)
J. Biol. Chem. 282, 5597-5607
   Abstract »    Full Text »    PDF »
Hyaluronan Constitutively Regulates Activation of Multiple Receptor Tyrosine Kinases in Epithelial and Carcinoma Cells.
S. Misra, B. P. Toole, and S. Ghatak (2006)
J. Biol. Chem. 281, 34936-34941
   Abstract »    Full Text »    PDF »
Transactivation of Sphingosine 1-Phosphate Receptors Is Essential for Vascular Barrier Regulation: NOVEL ROLE FOR HYALURONAN AND CD44 RECEPTOR FAMILY.
P. A. Singleton, S. M. Dudek, S.-F. Ma, and J. G. N. Garcia (2006)
J. Biol. Chem. 281, 34381-34393
   Abstract »    Full Text »    PDF »
Acylation of CD44 and Its Association with Lipid Rafts Are Required for Receptor and Hyaluronan Endocytosis.
S. P. Thankamony and W. Knudson (2006)
J. Biol. Chem. 281, 34601-34609
   Abstract »    Full Text »    PDF »
The Cannabinoid CB1 Receptor Antagonist Rimonabant (SR141716) Inhibits Human Breast Cancer Cell Proliferation through a Lipid Raft-Mediated Mechanism.
D. Sarnataro, S. Pisanti, A. Santoro, P. Gazzerro, A. M. Malfitano, C. Laezza, and M. Bifulco (2006)
Mol. Pharmacol. 70, 1298-1306
   Abstract »    Full Text »    PDF »
Tumor Cells Enhance Their Own CD44 Cleavage and Motility by Generating Hyaluronan Fragments.
K. N. Sugahara, T. Hirata, H. Hayasaka, R. Stern, T. Murai, and M. Miyasaka (2006)
J. Biol. Chem. 281, 5861-5868
   Abstract »    Full Text »    PDF »
Chemotaxis towards hyaluronan is dependent on CD44 expression and modulated by cell type variation in CD44-hyaluronan binding.
G. Tzircotis, R. F. Thorne, and C. M. Isacke (2005)
J. Cell Sci. 118, 5119-5128
   Abstract »    Full Text »    PDF »
Characterization of Promoter Elements of the Human HYAL-2 Gene.
G. Chow and W. Knudson (2005)
J. Biol. Chem. 280, 26904-26912
   Abstract »    Full Text »    PDF »
Antisense-Mediated Suppression of Hyaluronan Synthase 2 Inhibits the Tumorigenesis and Progression of Breast Cancer.
L. Udabage, G. R. Brownlee, M. Waltham, T. Blick, E. C. Walker, P. Heldin, S. K. Nilsson, E. W. Thompson, and T. J. Brown (2005)
Cancer Res. 65, 6139-6150
   Abstract »    Full Text »    PDF »
Protein Kinase A Gating of a Pseudopodial-located RhoA/ROCK/p38/NHE1 Signal Module Regulates Invasion in Breast Cancer Cell Lines.
R. A. Cardone, A. Bagorda, A. Bellizzi, G. Busco, L. Guerra, A. Paradiso, V. Casavola, M. Zaccolo, and S. J. Reshkin (2005)
Mol. Biol. Cell 16, 3117-3127
   Abstract »    Full Text »    PDF »
Regulation of MDR1 Expression and Drug Resistance by a Positive Feedback Loop Involving Hyaluronan, Phosphoinositide 3-Kinase, and ErbB2.
S. Misra, S. Ghatak, and B. P. Toole (2005)
J. Biol. Chem. 280, 20310-20315
   Abstract »    Full Text »    PDF »
Hyaluronan-CD44 Interaction with IQGAP1 Promotes Cdc42 and ERK Signaling, Leading to Actin Binding, Elk-1/Estrogen Receptor Transcriptional Activation, and Ovarian Cancer Progression.
L. Y. W. Bourguignon, E. Gilad, K. Rothman, and K. Peyrollier (2005)
J. Biol. Chem. 280, 11961-11972
   Abstract »    Full Text »    PDF »
Hyaluronan Constitutively Regulates ErbB2 Phosphorylation and Signaling Complex Formation in Carcinoma Cells.
S. Ghatak, S. Misra, and B. P. Toole (2005)
J. Biol. Chem. 280, 8875-8883
   Abstract »    Full Text »    PDF »
Myofibroblastic Differentiation Leads to Hyaluronan Accumulation through Reduced Hyaluronan Turnover.
R. H. Jenkins, G. J. Thomas, J. D. Williams, and R. Steadman (2004)
J. Biol. Chem. 279, 41453-41460
   Abstract »    Full Text »    PDF »

To Advertise     Find Products

Science Signaling. ISSN 1937-9145 (online), 1945-0877 (print). Pre-2008: Science's STKE. ISSN 1525-8882