Low Cholesterol Triggers Membrane Microdomain-dependent CD44 Shedding and Suppresses Tumor Cell Migration

Abstract

CD44 is a cell surface adhesion molecule for hyaluronan and is implicated in tumor invasion and metastasis. Proteolytic cleavage of CD44 plays a critical role in the migration of tumor cells and is regulated by factors present in the tumor microenvironment, such as hyaluronan oligosaccharides and epidermal growth factor. However, molecular mechanisms underlying the proteolytic cleavage on membranes remain poorly understood. In this study, we demonstrated that cholesterol depletion with methyl-␤-cyclodextrin, which disintegrates membrane lipid rafts, enhances CD44 shedding mediated by a disintegrin and metalloproteinase 10 (ADAM10) and that cholesterol depletion disorders CD44 localization to the lipid raft. We also evaluated the effect of long term cholesterol reduction using a statin agent and demonstrated that statin enhances CD44 shedding and suppresses tumor cell migration on a hyaluronan-coated substrate. Our results indicate that membrane lipid organization regulates CD44 shedding and propose a possible molecular mechanism by which cholesterol reduction might be effective for preventing and treating the progression of malignant tumors.CD44 is a cell surface receptor for several extracellular matrix components, including hyaluronan (1), and is implicated in a wide variety of biological processes, including cell migration (2) and tumor metastasis (3). The migratory properties of invasive tumor cells are affected both by the interaction between their adhesion molecules and the surrounding extracellular matrices and by growth factor signaling (4). The proteolytic ectodomain cleavage and release (shedding) of membrane proteins is a critical regulatory step in both physiological and pathological processes (5, 6).Recently, physiological inducers of CD44 shedding have been identified; hyaluronan small fragments, frequently detected in association with pathological conditions including cancer (7), induce CD44 shedding from tumor cells (8). Intracellular signaling via Rac GTPase, elicited by CD44 engagement, leads to ADAM 2 (a disintegrin and metalloproteinase)-mediated CD44 shedding and tumor cell migration (9). A prominent abnormality of tumor cells such as gliomas is the overexpression of the EGF receptor (10), and EGF induces the shedding of CD44 (11). Although the molecular mechanisms underlying CD44 shedding induced by spatio-temporally ordered intracellular signaling in the tumor microenvironment have been partly clarified (9, 11), the mechanism that triggers CD44 shedding on the membrane is not yet understood.Lipid rafts, which are microdomains of the plasma membrane that are rich in cholesterol and sphingolipids (12), are thought to exhibit a liquid-ordered phase floating in the liquid-disordered matrix of the plasma membrane. Recent studies have confirmed the importance of lipid rafts and their associated proteins, including CD44, in cancer progression (13). A number of reports have shown that CD44 is present in lipid rafts (14 -18). Recently, several reports have demonstrated that lipid rafts play a cruci...

Cholesterol lowering: role in cancer prevention and treatment

“…Treating human glioma cells with the lipid-raft-disrupting agent methyl-β-cyclodextrin (MβCD), which extracts cellular membrane cholesterol, results in increased CD44 shedding mediated by a disintegrin and metalloproteinase 10 (ADAM10) (Murai et al, 2011). Moreover, the CD44 shedding induced by cholesterol lowering is not limited to glioma cells, but is also seen in other tumor cells such as pancreatic cancer cells (Murai et al, 2011).…”

“…It was recently demonstrated that lipid rafts play a crucial role in the localization and functionality of CD44 (Murai et al, 2011) (Figure 2B). Treating human glioma cells with the lipid-raft-disrupting agent methyl-β-cyclodextrin (MβCD), which extracts cellular membrane cholesterol, results in increased CD44 shedding mediated by a disintegrin and metalloproteinase 10 (ADAM10) (Murai et al, 2011).…”

“…Treating human glioma cells with the lipid-raft-disrupting agent methyl-β-cyclodextrin (MβCD), which extracts cellular membrane cholesterol, results in increased CD44 shedding mediated by a disintegrin and metalloproteinase 10 (ADAM10) (Murai et al, 2011). Moreover, the CD44 shedding induced by cholesterol lowering is not limited to glioma cells, but is also seen in other tumor cells such as pancreatic cancer cells (Murai et al, 2011). CD44 shedding can also be induced by filipin, a polyene macrolide antibiotic that binds cholesterol and disperses it in the membrane, thereby disrupting lipid rafts by a different mechanism from MβCD (Murai et al, 2011).…”

Overexpression of sigma-1 receptor inhibits ADAM10 and ADAM17 mediated shedding in vitro

“…Cholesterol modification and sigma-1 receptor agonist DHEAS could reduce ionomycin-induced ADAM10-dependent BTC shedding but not ADAM17-dependent HB-EGF shedding Cholesterol is a ligand for sigma-1 receptor (Palmer et al, 2007) and its depletion can affect lipid raft formation and both ADAM10-dependent shedding (Murai et al, 2011) and ADAM17-dependent shedding (von Tresckow et al, 2004;Zimina et al, 2005;Tellier et al, 2006Tellier et al, , 2008. Therefore, we tested the effect of cholesterol in our system.…”

“…Those two transmembrane proteases are also involved in the shedding process of many other substrates, such as ligands of the epidermal growth factor receptor (EGFR), Notch1, L1 cell adhesion molecule (L1CAM), which are involved in pathological processes and human diseases such as cancer and stroke (Pruessmeyer and Ludwig, 2009). It is now known that lipid rafts can affect both ADAM17 and ADAM10 function as cholesterol depletion can trigger both ADAM10-mediated CD44 shedding (Murai et al, 2011) and ADAM17-mediated CD30 shedding (von Tresckow et al, 2004), collagen XVII shedding (Zimina et al, 2005), and TNF shedding (Tellier et al, 2006(Tellier et al, , 2008. It has been reported that ADAM10 is located in the nonraft region of the membrane of neuroblastoma SH-SY5Y cells when functioning as α-secretase of APP (Harris et al, 2009) and it cannot cleave APP in a cholesterolrich environment (Kojro et al, 2010).…”