Differential Activation of ERK and Rac Mediates the Proliferative and Anti-proliferative Effects of Hyaluronan and CD44 (2008)

Hyaluronan, a widely distributed component of the extracellular matrix, exists in a high molecular weight (native) form and lower molecular weight form (HMW- and LMW-HA, respectively). These different forms of hyaluronan bind to CD44 but elicit distinct effects on cellular function. A striking example is the opposing effects of HMW- and LMW-HA on the proliferation of vascular smooth muscle cells; the binding of HMW-HA to CD44 inhibits cell cycle progression, whereas the binding of LMW-HA to CD44 stimulates cell cycle progression. We now report that cyclin D1 is the primary target of LMW-HA in human vascular smooth muscle cells, as it is for HMW-HA, and that the opposing cell cycle effects of these CD44 ligands result from differential regulation of signaling pathways to cyclin D1. HMW-HA binding to CD44 selectively inhibits the GTP loading of Rac and Rac-dependent signaling to the cyclin D1 gene, whereas LMW-HA binding to CD44 selectively stimulates ERK activation and ERK-dependent cyclin D1 gene expression. These data describe a novel mechanism of growth control in which a ligand-receptor system generates opposing effects on mitogenesis by differentially regulating signaling pathways to a common cell cycle target. They also emphasize how a seemingly subtle change in matrix composition can have a profound effect on cell proliferation. Hyaluronan, a widely distributed component of the extracellular matrix, exists in a high molecular weight (native) form and lower molecular weight form (HMW- and LMW-HA, respectively). These different forms of hyaluronan bind to CD44 but elicit distinct effects on cellular function. A striking example is the opposing effects of HMW- and LMW-HA on the proliferation of vascular smooth muscle cells; the binding of HMW-HA to CD44 inhibits cell cycle progression, whereas the binding of LMW-HA to CD44 stimulates cell cycle progression. We now report that cyclin D1 is the primary target of LMW-HA in human vascular smooth muscle cells, as it is for HMW-HA, and that the opposing cell cycle effects of these CD44 ligands result from differential regulation of signaling pathways to cyclin D1. HMW-HA binding to CD44 selectively inhibits the GTP loading of Rac and Rac-dependent signaling to the cyclin D1 gene, whereas LMW-HA binding to CD44 selectively stimulates ERK activation and ERK-dependent cyclin D1 gene expression. These data describe a novel mechanism of growth control in which a ligand-receptor system generates opposing effects on mitogenesis by differentially regulating signaling pathways to a common cell cycle target. They also emphasize how a seemingly subtle change in matrix composition can have a profound effect on cell proliferation. The extracellular matrix (ECM) 2The abbreviations used are: ECM, extracellular matrix; cdk, cyclin-dependent kinase; DMEM, Dulbecco's modified Eagle's medium; FBS, fetal bovine serum; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HMW, high molecular weight (native) LMW, lower molecular weight; HA, hyaluronan; MMP, metalloproteinase; SMC, smooth muscle cell; Rb, retinoblastoma; siRNA, small interfering RNA; QPCR, quantitative PCR; ERK, extracellular signal-regulated kinase; BrdU, bromodeoxyuridine; MEF, mouse embryonic fibroblast; MAPK, mitogen-activated protein kinase; TGF-β, transforming growth factor-β. 2The abbreviations used are: ECM, extracellular matrix; cdk, cyclin-dependent kinase; DMEM, Dulbecco's modified Eagle's medium; FBS, fetal bovine serum; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HMW, high molecular weight (native) LMW, lower molecular weight; HA, hyaluronan; MMP, metalloproteinase; SMC, smooth muscle cell; Rb, retinoblastoma; siRNA, small interfering RNA; QPCR, quantitative PCR; ERK, extracellular signal-regulated kinase; BrdU, bromodeoxyuridine; MEF, mouse embryonic fibroblast; MAPK, mitogen-activated protein kinase; TGF-β, transforming growth factor-β. is a key regulator of several cellular functions, including differentiation, survival, and proliferation. Hyaluronan (HA) is a widely distributed component of the ECM (1Fraser J.R. Laurent T.C. Laurent U.B. J. Intern. Med. 1997; 242: 27-33Crossref PubMed Scopus (1457) Google Scholar, 2Noble P.W. Matrix Biol. 2002; 21: 25-29Crossref PubMed Scopus (461) Google Scholar). Unlike ECM proteins such as collagen, fibronectin, vitronectin, and laminin, HA is a glycosaminoglycan with a molecular mass ranging from several million daltons (called native or high molecular weight HA (HMW-HA)) to 20–500 kDa (called lower molecular weight HA (LMW-HA)). HMW-HA is produced and secreted from several cell types where it is incorporated into the ECM and interacts in an autocrine and paracrine fashion with its receptors. LMW-HA can be synthesized de novo or generated from the enzymatic degradation or oxidative hydrolysis of HMW-HA, typically at sites of inflammation (2Noble P.W. Matrix Biol. 2002; 21: 25-29Crossref PubMed Scopus (461) Google Scholar, 3Hawkins C.L. Davies M.J. Free Radic. Biol. Med. 1998; 24: 1396-1410Crossref PubMed Scopus (108) Google Scholar). Very small HA oligomers also have biological activities (4Ghatak S. Misra S. Toole B.P. J. Biol. Chem. 2002; 277: 38013-38020Abstract Full Text Full Text PDF PubMed Scopus (306) Google Scholar, 5Termeer C.C. Hennies J. Voith U. Ahrens T. Weiss J.M. Prehm P. Simon J.C. J. Immunol. 2000; 165: 1863-1870Crossref PubMed Scopus (331) Google Scholar), but their biological significance is less well understood. HA can bind to several cell surface receptors (6Hardwick C. Hoare K. Owens R. Hohn H.P. Hook M. Moore D. Cripps V. Austen L. Nance D.M. Turley E.A. J. Cell Biol. 1992; 117: 1343-1350Crossref PubMed Scopus (320) Google Scholar, 7Bono P. Rubin K. Higgins J.M. Hynes R.O. Mol. Biol. Cell. 2001; 12: 891-900Crossref PubMed Scopus (115) Google Scholar), but the most extensively studied receptor for HA is CD44 (8Aruffo A. Stamenkovic I. Melnick M. Underhill C.B. Seed B. 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Sequential activation of the metalloproteinase, MMP-1, and presenilin-dependent γ-secretase releases a biologically active extracellular domain and intracellular domain from CD44, which appears to explain some of the effects of CD44 on gene transcription (13Nagano O. Saya H. Cancer Sci. 2004; 95: 930-935Crossref PubMed Scopus (310) Google Scholar). CD44 plays an important role in regulation of the cortical actin cytoskeleton through an interaction with merlin and ERM proteins and also transmits signals that control differentiation, migration, and proliferation (14Yonemura S. Hirao M. Doi Y. Takahashi N. Kondo T. Tsukita S. J. Cell Biol. 1998; 140: 885-895Crossref PubMed Scopus (504) Google Scholar, 15Bretscher A. Edwards K. Fehon R.G. Nat. Rev. Mol. Cell Biol. 2002; 3: 586-599Crossref PubMed Scopus (1127) Google Scholar). In addition to HA, CD44 binds to other ligands, including osteopontin and MMPs (16Weber J.D. Raben D.M. Philips P.J. Baldassare J.J. Biochem. J. 1997; 326: 61-68Crossref PubMed Scopus (375) Google Scholar, 17Johnson C. Galis Z.S. Arterioscler. Thromb. Vasc. Biol. 2004; 24: 54-60Crossref PubMed Scopus (274) Google Scholar). In fact, the binding of MMPs to CD44 promotes the MMP-dependent activation of TGF-β (18Yu Q. Stamenkovic I. Genes Dev. 2000; 14: 163-176PubMed Google Scholar). Thus, the signaling effects of CD44 can be direct, a consequence of cortical actin organization, or indirect through TGF-β. Both HMW-HA and LMW-HA bind to CD44 with similar affinities, but the biological outcome of the binding is distinct (10Puré E. Cuff C.A. Trends Mol. Med. 2001; 7: 213-221Abstract Full Text Full Text PDF PubMed Scopus (357) Google Scholar). For example, the binding of HMW-HA to CD44 antagonizes mitogen-induced cell cycle progression, whereas the binding of LMW-HA to CD44 stimulates cell cycle progression in vascular smooth muscle cells (SMCs) and fibroblasts (19Cuff C.A. Kothapalli D. Azonobi I. Chun S. Zhang Y. Belkin R. Yeh C. Secreto A. Assoian R.K. Rader D.J. Puré E. J. Clin. Investig. 2001; 108: 1031-1040Crossref PubMed Scopus (255) Google Scholar). The molecular basis for these opposing proliferative effects of HMW-HA and LMW-HA remains unresolved. We recently reported that the anti-proliferative effect of HMW-HA reflects its ability to inhibit the mitogen-dependent induction of cyclin D1 mRNA and protein (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar). This effect is mediated by CD44 and likely independent of TGF-β activation because the anti-proliferative effects of TGF-β are most closely linked to up-regulation of cyclin-dependent kinase (cdk) inhibitors, p15INK4b and p21cip1 (21Datto M.B. Hu P.P. Kowalik T.F. Yingling J. Wang X.F. Mol. Cell. Biol. 1997; 17: 2030-2037Crossref PubMed Google Scholar). The signal transduction mechanism(s) by which HMW-HA binding to CD44 inhibits cyclin D1 gene expression remains unexplored. The cyclin D1 gene is the target of many mitogenic signaling pathways, but the best studied pathway to cyclin D1 is arguably the ERK-MAPK pathway. In several cell types, a sustained ERK activity between 3 and 6 h after mitogen stimulation results in the induction of cyclin D1 mRNA in mid-G1 phase, at least when cells are re-entering G1 phase from quiescence (22Balmanno K. Cook S.J. Oncogene. 1999; 18: 3085-3097Crossref PubMed Scopus (201) Google Scholar, 23Welsh C.F. Roovers K. Villanueva J. Liu Y. Schwartz M.A. Assoian R.K. Nat. Cell Biol. 2001; 3: 950-957Crossref PubMed Scopus (269) Google Scholar). A second common way to induce cyclin D1 mRNA is through Rac (24Westwick J.K. Lambert Q.T. Clark G.J. Symons M. Van Aelst L. Pestell R.G. Der C.J. Mol. Cell. Biol. 1997; 17: 1324-1335Crossref PubMed Scopus (385) Google Scholar, 25Klein E.A. Yang C. Kazanietz M.G. Assoian R.K. Cell Cycle. 2007; 6: 1115-1121Crossref PubMed Scopus (21) Google Scholar). Rac signaling to cyclin D1 requires NF-κB and results in a somewhat earlier induction of cyclin D1 within the G1 phase (23Welsh C.F. Roovers K. Villanueva J. Liu Y. Schwartz M.A. Assoian R.K. Nat. Cell Biol. 2001; 3: 950-957Crossref PubMed Scopus (269) Google Scholar, 26Fournier A.K. Campbell L.E. Castagnino P. Liu W.F. Chung B.M. Weaver V.M. Chen C.S. Assoian R.K. J. Cell Sci. 2008; 121: 226-233Crossref PubMed Scopus (51) Google Scholar). Once expressed, cyclin D1 binds to cdk4 or cdk6 and initiates the inactivating phosphorylation of Rb that, in turn, controls the transcription of E2F-dependent genes and progression into S phase (27Sherr C.J. Roberts J.M. Genes Dev. 1999; 13: 1501-1512Crossref PubMed Scopus (5108) Google Scholar, 28Dimova D.K. Dyson N.J. Oncogene. 2005; 24: 2810-2826Crossref PubMed Scopus (586) Google Scholar). In this study we show that the opposing effects of HMW-HA and LMW-HA on S phase entry do not reflect regulation of distinct cell cycle targets but rather opposing effects on the expression of cyclin D1 mRNA. Moreover, we demonstrate that HMW-HA inhibits cyclin D1 gene expression by selectively inhibiting Rac signaling to cyclin D1, whereas LMW-HA stimulates cyclin D1 gene expression by selectively stimulating ERK signaling to cyclin D1. Both effects require CD44. To our knowledge, this is the first study of a ligand-receptor system having opposing effects on mitogenesis by differentially regulating signaling pathways to a common cell cycle target. Cell Culture—Early passage human vascular SMCs were maintained and serum-starved at near confluence for 48 h as described (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar). Early passage vascular SMCs were isolated from wild-type and CD44-null C57Bl/6 mice and cultured as described (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar). Primary mouse embryonic fibroblasts (MEFs) were isolated from E14.5 background-matched wild-type or K. H. M. I. K. M. R. T. N. M. S. J. 2000; PubMed Scopus Google Scholar). The were maintained in and used at passage were serum-starved in DMEM, bovine for 48 A of LMW-HA or HMW-HA to with the were with or for with HMW-HA or LMW-HA, and to the serum-starved S phase entry by as described (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar, E.A. Y. Castagnino P. Kothapalli D. Assoian R.K. 2007; PubMed Scopus Google Scholar) in cells with from the of typically to the cells as described (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar), similar results were in the or of B. In some a to human CD44 or an to the the addition of The show typically from 3 to independent and were as described (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar) human vascular SMCs in The Rac or a The cells were with with in the or of LMW-HA For human vascular SMCs were and cells in DMEM, The and were in and to the cells for h the cells for an h in E.A. Y. Castagnino P. Kothapalli D. Assoian R.K. 2007; PubMed Scopus Google Scholar). The cells were with in the or of LMW-HA for The used to cyclin D1 and have described E.A. Y. Castagnino P. Kothapalli D. Assoian R.K. 2007; PubMed Scopus Google Scholar). Rac used A used as control Rac SMCs cells were serum-starved for 48 The cells were with HMW-HA or LMW-HA for at for 3 or with or FBS, Rac activity a Rac were and as the of with were in and The of cyclin D1 cdk4 and S were by as described (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar, E.A. Y. Castagnino P. Kothapalli D. Assoian R.K. 2007; PubMed Scopus Google Scholar). results for cyclin D1 and were to S or cdk4 of which in to the used of and the proteins were to with to cyclin D1, Rb, and actin as described (23Welsh C.F. Roovers K. Villanueva J. Liu Y. Schwartz M.A. Assoian R.K. Nat. Cell Biol. 2001; 3: 950-957Crossref PubMed Scopus (269) Google Scholar, J. Y. Assoian R.K. Mol. Biol. Cell. 2007; 18: PubMed Scopus Google Scholar, Kothapalli D. Y. Assoian R.K. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus (21) Google Scholar). The proteins were were by and were LMW-HA with a to S reported that LMW-HA stimulates S phase entry in mouse vascular SMCs in FBS, and that the effect requires CD44 (23Welsh C.F. Roovers K. Villanueva J. Liu Y. Schwartz M.A. Assoian R.K. Nat. Cell Biol. 2001; 3: 950-957Crossref PubMed Scopus (269) Google Scholar). in the of vascular SMCs a of the We the effect of LMW-HA, and the on CD44, in human vascular SMCs A and These also that the of LMW-HA similar to LMW-HA not S phase in the of that LMW-HA binding to CD44 promotes mitogenesis by with a mitogenic in cellular or of vascular SMCs in the or of or LMW-HA D1 the Primary of the effect of LMW-HA on G1 phase cell cycle progression in human vascular The of Rb is a G1 phase that is mediated by and for the of and is for S phase entry D.K. Dyson N.J. Oncogene. 2005; 24: 2810-2826Crossref PubMed Scopus (586) Google Scholar, C. A. Oncogene. PubMed Scopus Google Scholar). LMW-HA Rb as by it also the cyclin A an target J. Kowalik T. J.R. Mol. Cell. Biol. PubMed Scopus Google Scholar, J.M. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar), and cyclin A mRNA We the effect of LMW-HA within the G1 phase and that LMW-HA the expression of cyclin D1 mRNA and protein and respectively). LMW-HA also the protein of the but not the expression of mRNA of the G1 phase cdk4 and were by LMW-HA were to in these cells not these data that the of Rb and S phase entry in to LMW-HA results from up-regulation of cyclin D1, of and the activation of cdk4 and The of are typically by through the E. A. M. Nat. Cell Biol. 1999; PubMed Scopus Google Scholar, T. M. Nat. Rev. Mol. Cell Biol. 2004; PubMed Scopus Google Scholar). is the component of the and its mRNA and protein are by as cells S phase from quiescence Y. Roberts J.M. Assoian R.K. J. Cell Biol. 2007; PubMed Scopus Google Scholar). with these we that LMW-HA the expression of mRNA and protein A the effects of LMW-HA on cyclin D1 and and respectively). LMW-HA to the expression of cyclin D1 and in CD44-null SMCs and Thus, the effects of LMW-HA on cyclin D1 and require CD44. We recently reported that the of in to mitogenic stimulation can be a consequence of cyclin D1 induction and of Rb promotes which and J. Y. Assoian R.K. Mol. Biol. Cell. 2007; 18: PubMed Scopus Google Scholar, Y. Roberts J.M. Assoian R.K. J. Cell Biol. 2007; PubMed Scopus Google Scholar). we that the effects of LMW-HA on and were to the induction of cyclin D1 because cyclin D1 with the effect of LMW-HA on gene induction and In of not LMW-HA from cyclin D1 mRNA as be up-regulation a effect of these data that cyclin D1 is the primary target of our report that HMW-HA inhibits S phase entry by the mitogen-dependent induction of cyclin D1 (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar), we that the opposing effect of HMW-HA and LMW-HA are through a common cell cycle cyclin D1. with this of cyclin D1 S phase entry in to LMW-HA This result also that cyclin D1 is the cyclin regulating S phase entry in this of HMW- and LMW-HA on and Rac-dependent D1 ERK-MAPK and Rac pathways have linked to the induction of cyclin D1 mRNA We the effect of these pathways on cyclin D1 gene expression in human vascular SMCs and these pathways be differentially by HMW-HA and We that induction of cyclin D1 mRNA and protein were not by ERK activation with the In expression of Rac a or distinct cyclin D1 gene expression in human vascular SMCs and also the induction of cyclin D1 protein We HMW-HA and LMW-HA differentially ERK or Rac signaling to cyclin D1 mRNA. We sustained ERK activity h after mitogenic because that signaling linked to the induction of cyclin D1 mRNA We on earlier for Rac activation because the GTP loading of Rac is to We that the effect of HMW-HA on cyclin D1 gene expression not with a in ERK activation HMW-HA GTP loading of Rac in human vascular SMCs the effect of LMW-HA on cyclin D1 gene expression to not with a stimulation of loading Rac inhibit the induction of cyclin D1 mRNA or S phase entry in cells LMW-HA result in the activation of by for in vascular SMCs with the effects of LMW-HA on ERK activity and cyclin D1 induction are linked because with the induction of cyclin D1 mRNA in to LMW-HA these data show that HMW- and LMW-HA their opposing effects on cyclin D1 gene expression by differentially regulating the activation of Rac and we that HA binding to CD44 can have opposing effects on mitogenesis because the high and lower molecular weight forms of HA differentially regulate the and pathways to cyclin D1 of LMW-HA on ERK and Rac signaling to cyclin D1 human vascular SMCs were with a or distinct with and LMW-HA, or FBS, in the of cells were and for Rac and and used to of cyclin D1 mRNA and S at h after human vascular SMCs were with for in the or of cells were and for and human vascular SMCs were with and LMW-HA in the of or from some cells and used to of cyclin D1 mRNA and S by The expression of cyclin D1 mRNA to S The results described the mechanism the effect of LMW-HA, and to explain how HMW-HA and LMW-HA can have opposing effects on cell cycle progression binding to the CD44. We show that LMW-HA stimulates the induction of cyclin D1 in cells to a mitogenic LMW-HA also stimulates the expression of and the of but which we cyclin D1 or that the effects on and likely result from cyclin Rb Thus, the effect of LMW-HA on cyclin D1 is primary and its effects on and are is that HMW-HA its effect by inhibiting mitogen-dependent cyclin D1 induction in the cells (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar). Thus, HMW-HA and LMW-HA have opposing effects on the expression of cyclin D1, and in the HA effects require CD44 study and C.A. Kothapalli D. Azonobi I. Chun S. Zhang Y. Belkin R. Yeh C. Secreto A. Assoian R.K. Rader D.J. Puré E. J. Clin. Investig. 2001; 108: 1031-1040Crossref PubMed Scopus (255) Google Scholar, D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar). These results emphasize how a seemingly subtle change in matrix the of HA, can result in distinct effects on cell proliferation. that CD44 can with in its signals D. J. P.W. Rev. Cell Dev. Biol. 2007; PubMed Scopus Google Scholar), but with other K. N. S. R. T. Takahashi M. K. S. M. 2007; PubMed Scopus Google Scholar, Immunol. 2008; PubMed Scopus Google Scholar), we were to surface expression by in human vascular smooth muscle cells or with HMW-HA or LMW-HA, to HMW-HA or LMW-HA have effect on the surface of CD44 not The ERK-MAPK pathway and the Rac pathway cyclin D1 gene expression in several cell types, and we show that pathways to the mitogen-dependent induction of cyclin D1 mRNA in human vascular ERK and Rac activities are in to several receptor types, including receptor and signaling by CD44 is not well the by which CD44 is to signal regulation of ERM proteins and of its intracellular are different from the with activation of receptor and Thus, it appears that a of signaling to regulate the activation of and of the in this study is that the opposing effects of HMW-HA and LMW-HA on cyclin D1 gene expression result from differential regulation of the ERK and Rac pathways to cyclin D1 mRNA the effect of HMW-HA results from of mitogen-dependent loading Rac signaling to cyclin whereas the effect of LMW-HA results from a of ERK activation in to mitogenic We do not how the binding of HMW-HA LMW-HA to CD44 differentially regulate the ERK and Rac pathways, but we that it differential of ERM proteins or merlin have linked to ERK and as well as differential binding of CD44 to a receptor kinase or M. A. I. V. N. Der C.J. T. Dev. Cell. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, H. T. J. M. H. Herrlich P. Cancer 2007; PubMed Scopus Google Scholar). with the we that have the effect of CD44 on growth to an between CD44 and the growth receptor L. P. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, L. T. B. P. Biochem. J. 2007; PubMed Scopus Google Scholar). In HA exists in a between its high molecular weight form and lower molecular forms that from the of or from oxidative degradation The degradation of HMW-HA to LMW-HA at sites of inflammation in and and we have (20Kothapalli D. Zhao L. Hawthorne E.A. Cheng Y. Lee E. Puré E. Assoian R.K. J. Cell Biol. 2007; 176: 535-544Crossref PubMed Scopus (66) Google Scholar) that this HA and CD44 to as a proliferation in the of HMW-HA the effect of CD44. the in the of LMW-HA the effect of CD44. This study that the by differentially regulating the ERK and Rac signaling pathways to a common cell cycle cyclin D1 our on cell cycle progression, it is that ERK and Rac regulate cell and as well as proliferation A. A. PubMed Scopus Google Scholar, J. Cell Biol. 2000; PubMed Scopus Google Scholar). of CD44 to ERK and Rac by HMW- and LMW-HA have for the and effects of CD44. We and for We also and Kazanietz for with