Hyaluronan-mediated CD44 activation of RhoGTPase signaling and cytoskeleton function promotes tumor progression

doi:10.1016/j.semcancer.2008.03.007

Seminars in Cancer Biology

Volume 18, Issue 4, August 2008, Pages 251-259

https://doi.org/10.1016/j.semcancer.2008.03.007 Get rights and content

Abstract

Hyaluronan (HA), a major component of the extracellular matrix (ECM), is enriched in many types of tumors. In cancer patients HA concentrations are usually higher in malignant tumors than in corresponding benign or normal tissues, and in some tumor types the level of HA is predictive of malignancy. HA is often bound to CD44 isoforms which are ubiquitous, abundant, and functionally important cell surface receptors. This article reviews the current evidence for HA/CD44-mediated activation of the ankyrin-based cytoskeleton and RhoGTPase signaling during tumor progression. A special focus is placed on the role of HA-mediated CD44 interaction with unique downstream effectors (e.g., the cytoskeletal protein, ankyrin and/or various GTPases (e.g., RhoA, Rac1 and Cdc42)) in coordinating intracellular signaling pathways (e.g., Ca²⁺ mobilization, Rho signaling, PI3 kinase-AKT activation, NHE1-mediated cellular acidification, transcriptional upregulation and cytoskeletal function) and generating the concomitant onset of tumor cell activities (e.g., tumor cell adhesion, growth, survival, migration and invasion) and tumor progression. I believe this information will provide valuable new insights into poorly understood aspects of solid tumor malignancy. Furthermore, the new knowledge concerning HA/CD44-mediated oncogenic signaling events will have potentially important clinical utility, and could establish CD44 and its associated signaling molecules as important tumor markers for the early detection and evaluation of oncogenic potential. It could also serve as ground work for the future development of new drug targets to inhibit HA/CD44-mediated tumor metastasis and cancer progression.

Section snippets

Hyaluronan (HA) and CD44 in tumor progression

Tumor invasion and metastasis are the primary causes of morbidity in patients diagnosed with solid tumors such as breast cancer [1], ovarian cancer [2] and squamous cell carcinomas (SCC) [3]. It is now certain that both oncogenic signaling and cytoskeleton functions are directly involved in tumor cell growth, migration, invasion of surrounding tissue, and metastasis [4], [5]. A number of studies have aimed at identifying these molecules which are specifically expressed by epithelial tumor cells

HA–CD44 interaction with the cytoskeletal protein, ankyrin

CD44 interacts with a number of membrane-associated cytoskeletal proteins, such as ankyrin, which are expressed in a variety of biological systems [14], [34]. Ankyrin contains three functional domains: a conserved N-terminal ankyrin repeat domain (ARD) (consisting of 22–24 tandem repeats of 33 amino acids with a consensus sequence such as -G-TPLH-AA--GH---V/A--LL--GA--ND---); a spectrin binding domain; and a variably sized C-terminal regulatory domain [35] (Fig. 1). CD44 binds directly to the

HA/CD44-mediated RhoGTPase signaling in regulating tumor progression

RhoGTPases (e.g., RhoA, Rac1 and Cdc42) belong to members of the Rho subclass of the Ras superfamily [47], and are known to cycle between an active GTP-bound state and an inactive GDP-bound state to transmit diverse signals from cell surface receptors to intracellular targets. They function as molecular switches that, in response to external stimuli, regulate key signaling pathways and control a variety of cellular activities (Fig. 2) including gene transcription, cytoskeleton reorganization,

Summary

HA/CD44-mediated tumor cell-specific phenotypes are closely linked to cytoskeletal functions which involve membrane-associated cytoskeletal proteins (e.g., ankyrin) and the small GTP-binding proteins such as RhoA, Rac1 and Cdc42. Activation of ankyrin and RhoGTPases (e.g., Rac1, Cdc42 and RhoA) has been shown to produce specific structural changes in actin assembly, cytoskeleton reorganization, transcriptional activation, tumor cell growth, survival, migration and invasion. In summary, a

Acknowledgements

I gratefully acknowledge Dr. Gerard J. Bourguignon's assistance in the preparation of this paper. I am also grateful for Ms. Christine Camacho for her help in preparing graphs and illustrations. This work was supported by United States Public Health grants (R01 CA66163, R01 CA78633 and P01 AR39448), a VA Merit Review grant and a DOD grant. L.Y.W.B is a VA Research Career Scientist.

References (105)

D.A. Lauffenburger et al.
Cell migration: a physically integrated molecular process
Cell
(1996)
J.Y. Lee et al.
Hyaluronan: a multifunctional, megadalton, stealth molecule
Curr Opin Cell Biol
(2000)
B.P. Toole
Hyaluronan in morphogenesis
Semin Cell Dev Biol
(2001)
B.P. Toole et al.
Hyaluronan-cell interactions in cancer and vascular disease
J Biol Chem
(2002)
L.Y.W. Bourguignon et al.
Interaction between CD44 and p185^neu promotes human ovarian tumor cell activation
J Biol Chem
(1997)
U. Gunthert et al.
A new variant of glycoprotein CD44 confers metastatic potential to rat carcinoma cells
Cell
(1991)
E.A. Turley et al.
Signaling properties of hyaluronan receptors
J Biol Chem
(2002)
L.Y.W. Bourguignon et al.
Hyaluronan-mediated CD44 interaction with RhoGEF and Rho-kinase promotes Grb2-associated binding-1 phosphorylation and phosphatidylinositol 3-kinase signaling leading to cytokine (macrophage-colony stimulating factor) production and breast tumor progression
J Biol Chem
(2003)
L.Y.W. Bourguignon et al.
CD44 interaction with Tiam1 promotes Rac1 signaling and hyaluronic acid (HA)-mediated breast tumor cell migration
J Biol Chem
(2000)
L.Y.W. Bourguignon et al.
Hyaluronan (HA) promotes CD44v3–Vav2 interaction with Grb2–p185^HER2 and induces Rac1 & Ras signaling during ovarian tumor cell migration and growth
J Biol Chem
(2001)

L.Y.W. Bourguignon et al.

CD44 interaction with c-Src kinase promotes cortactin-mediated cytoskeleton function and hyaluronic acid (HA)-dependent ovarian tumor cell migration

J Biol Chem

(2001)

L.Y.W. Bourguignon et al.

Hyaluronan (HA) promotes signaling interaction between CD44 and the TGF-RI receptor in metastatic breast tumor cells

J Biol Chem

(2002)

G.R. Screaton et al.

The identification of a new alternative exon with highly restricted tissue expression in transcripts encoding the mouse Pgp-1 (CD44) homing receptor. Comparison of all 10 variable exons between mouse, human and rat

J Biol Chem

(1993)

V.B. Lokeshwar et al.

The cell adhesion molecule, GP116, is a new CD44 variant (ex14/v10) involved in hyaluronic acid binding and endothelial cell proliferation

J Biol Chem

(1996)

P.A. Singleton et al.

CD44 interaction with ankyrin and IP₃ receptors in lipid rafts promotes hyaluronan-mediated Ca²⁺ signaling leading to nitric oxide production and endothelial cell adhesion and proliferation

Exp Cell Res

(2004)

L.Y. Bourguignon et al.

The involvement of the cytoskeleton in regulating IP3 receptor-mediated internal Ca²⁺ release in human blood platelets

Cell Biol Int

(1993)

L.Y. Bourguignon et al.

The involvement of ankyrin in the regulation of inositol 1,4,5-trisphosphate receptor-mediated internal Ca²⁺ release from Ca²⁺ storage vesicles in mouse T-lymphoma cells

J Biol Chem

(1993)

L.Y. Bourguignon et al.

Identification of the ankyrin-binding domain of the mouse T-lymphoma cell inositol 1,4,5-trisphosphate (IP3) receptor and its role in the regulation of IP3-mediated internal Ca²⁺ release

J Biol Chem

(1995)

Y. Zheng et al.

Guanine nucleotide exchange catalyzed by dbl oncogene product

Methods Enzymol

(1995)

S. Fukuhara et al.

A novel PDZ domain containing guanine nucleotide exchange factor links heterotrimeric G proteins to Rho

J Biol Chem

(1999)

M.A. Lemmon et al.

Pleckstrin homology domains and the cytoskeleton

FEBS Lett

(2002)

M. Amano et al.

Phosphorylation and activation of myosin by Rho-associated kinase (Rho-kinase)

J Biol Chem

(1996)

Y. Kawano et al.

Ras oncoprotein induces CD44 cleavage through phosphoinositide 3-OH kinase and the rho family of small G proteins

J Biol Chem

(2000)

L. Counillon et al.

The expanding family of eucaryotic Na(+)/H(+) exchangers

J Biol Chem

(2000)

S.J. Reshkin et al.

Phosphoinositide 3-kinase is involved in the tumor-specific activation of human breast cancer cell Na(+)/H(+) exchange, motility, and invasion induced by serum deprivation

J Biol Chem

(2000)

L.Y. Bourguignon et al.

CD44 interaction with Na+–H+ exchanger (NHE1) creates acidic microenvironments leading to hyaluronidase-2 and cathepsin B activation and breast tumor cell invasion

J Biol Chem

(2004)

H. Chikumi et al.

Regulation of G protein-linked guanine nucleotide exchange factors for Rho, PDZ-RhoGEF, and LARG by tyrosine phosphorylation: evidence of a role for focal adhesion kinase

J Biol Chem

(2002)

L.Y. Bourguignon et al.

Hyaluronan–CD44 interaction with leukemia-associated RhoGEF and epidermal growth factor receptor promotes Rho/Ras co-activation, phospholipase C epsilon-Ca²⁺ signaling, and cytoskeleton modification in head and neck squamous cell carcinoma cells

J Biol Chem

(2006)

A.J. Ridley et al.

The small GTP-binding protein rac regulates growth factor-induced membrane ruffling

Cell

(1992)

I.N. Fleming et al.

Lysophosphatidic acid induces threonine phosphorylation of Tiam1 in Swiss 3T3 fibroblasts via activation of protein kinase C

J Biol Chem

(1997)

I.N. Fleming et al.

Ca²⁺/calmodulin-dependent protein kinase II regulates Tiam1 by reversible protein phosphorylation

J Biol Chem

(1999)

L.Y. Bourguignon et al.

Heregulin-mediated ErbB2–ERK signaling activates hyaluronan synthases leading to CD44-dependent ovarian tumor cell growth and migration

J Biol Chem

(2007)

M.W. Briggs et al.

IQGAP1 as signal integrator: Ca²⁺, calmodulin, Cdc42 and the cytoskeleton

FEBS Lett

(2003)

J.M. Mataraza et al.

IQGAP1 promotes cell motility and invasion

J Biol Chem

(2003)

M. Roy et al.

IQGAP1 binds ERK2 and modulates its activity

J Biol Chem

(2004)

B. Parker et al.

Distant metastasis in breast cancer: molecular mechanisms and therapeutic targets

Cancer Biol Ther

(2003)

S. Bhoola et al.

Diagnosis and management of epithelial ovarian cancer

Obstet Gynecol

(2006)

R.H. Kramer et al.

Tumor cell invasion and survival in head and neck cancer

Cancer Metastasis Rev

(2005)

W.G. Jiang et al.

Molecular and cellular basis of cancer invasion and metastasis: implications for treatment

Br J Surg

(1994)

T.C. Laurent et al.

Hyaluronan

FASEB J

(1992)

R. Stern et al.

Hyaluronidases: their genomics, structures, and mechanisms of action

Chem Rev

(2006)

W. Knudson et al.

The role and regulation of tumor-associated hyaluronan

Ciba Found Symp

(1989)

B. Delpech et al.

Serum hyaluronan (hyaluronic acid) in breast cancer patients

Int J Cancer

(1990)

N. Iida et al.

New CD44 splice variants associated with human breast cancers

J Cell Physiol

(1995)

D. Zhu et al.

The ankyrin-binding domain of CD44s is involved in regulating hyaluronic acid-mediated functions and prostate tumor cell transformation

Cell Motil Cytoskeleton

(1998)

N. Iida et al.

Coexpression of CD44 variant (v10/ex 14) and CD44s in human mammary epithelial cells promotes tumorinogenesis

J Cell Physiol

(1997)

P. Auvinen et al.

Expression of CD44s, CD44v3 and CD44v6 in benign and malignant breast lesions: correlation and colocalization with hyaluronan

Histopathology

(2005)

L.Y.W. Bourguignon

CD44-mediated oncogenic signaling and cytoskeleton activation during mammary tumor progression

J Mammary Gland Biol Neoplasia

(2001)

L.Y.W. Bourguignon et al.

CD44 Isoform-cytoskeleton interaction in oncogenic signaling and tumor progression

Front Biosci

(1998)

L.Y.W. Bourguignon et al.

CD44v_3,8–10-cytoskeleton interaction is involved in matrix metalloproteinase (MMP-9) function and tumor cell migration and invasion in metastatic breast tumor cells

J Cell Physiol

(1998)

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ReviewHyaluronan-mediated CD44 activation of RhoGTPase signaling and cytoskeleton function promotes tumor progression

Abstract

Section snippets

Hyaluronan (HA) and CD44 in tumor progression

HA–CD44 interaction with the cytoskeletal protein, ankyrin

HA/CD44-mediated RhoGTPase signaling in regulating tumor progression

Summary

Acknowledgements

Cell

Curr Opin Cell Biol

Semin Cell Dev Biol

J Biol Chem

J Biol Chem

Cell

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

Exp Cell Res

Cell Biol Int

J Biol Chem

J Biol Chem

Methods Enzymol

J Biol Chem

FEBS Lett

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

Cell

J Biol Chem

J Biol Chem

J Biol Chem

FEBS Lett

J Biol Chem

J Biol Chem

Distant metastasis in breast cancer: molecular mechanisms and therapeutic targets

Cancer Biol Ther

Diagnosis and management of epithelial ovarian cancer

Obstet Gynecol

Tumor cell invasion and survival in head and neck cancer

Cancer Metastasis Rev

Molecular and cellular basis of cancer invasion and metastasis: implications for treatment

Br J Surg

Hyaluronan

FASEB J

Hyaluronidases: their genomics, structures, and mechanisms of action

Chem Rev

The role and regulation of tumor-associated hyaluronan

Ciba Found Symp

Serum hyaluronan (hyaluronic acid) in breast cancer patients

Int J Cancer

New CD44 splice variants associated with human breast cancers

J Cell Physiol

The ankyrin-binding domain of CD44s is involved in regulating hyaluronic acid-mediated functions and prostate tumor cell transformation

Cell Motil Cytoskeleton

Coexpression of CD44 variant (v10/ex 14) and CD44s in human mammary epithelial cells promotes tumorinogenesis

J Cell Physiol

Expression of CD44s, CD44v3 and CD44v6 in benign and malignant breast lesions: correlation and colocalization with hyaluronan

Histopathology

CD44-mediated oncogenic signaling and cytoskeleton activation during mammary tumor progression

J Mammary Gland Biol Neoplasia

CD44 Isoform-cytoskeleton interaction in oncogenic signaling and tumor progression

Front Biosci

CD44v3,8–10-cytoskeleton interaction is involved in matrix metalloproteinase (MMP-9) function and tumor cell migration and invasion in metastatic breast tumor cells

J Cell Physiol

Review
Hyaluronan-mediated CD44 activation of RhoGTPase signaling and cytoskeleton function promotes tumor progression

CD44v_3,8–10-cytoskeleton interaction is involved in matrix metalloproteinase (MMP-9) function and tumor cell migration and invasion in metastatic breast tumor cells