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 Jun 25;279(26):26991-7007. doi: 10.1074/jbc.M311838200. Epub 2004 Apr 16.

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.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amides / pharmacology
  • Amiloride / analogs & derivatives*
  • Amiloride / pharmacology
  • Animals
  • Breast Neoplasms / enzymology
  • Breast Neoplasms / metabolism*
  • Cathepsin B / metabolism*
  • Caveolin 1
  • Caveolins / metabolism
  • Cell Line, Tumor
  • Endosomes / metabolism
  • Enzyme Inhibitors / pharmacology
  • Humans
  • Hyaluronan Receptors / chemistry
  • Hyaluronan Receptors / genetics
  • Hyaluronan Receptors / metabolism*
  • Hyaluronic Acid / metabolism
  • Hyaluronoglucosaminidase / genetics
  • Hyaluronoglucosaminidase / metabolism*
  • Hydrogen-Ion Concentration
  • Immunoglobulin G / pharmacology
  • Intracellular Signaling Peptides and Proteins
  • Lysosomes / metabolism
  • Membrane Microdomains / chemistry
  • Membrane Microdomains / metabolism*
  • Membrane Proteins / metabolism
  • Neoplasm Invasiveness
  • Phosphorylation
  • Protein Serine-Threonine Kinases / antagonists & inhibitors
  • Protein Serine-Threonine Kinases / chemistry
  • Protein Serine-Threonine Kinases / metabolism
  • Pyridines / pharmacology
  • RNA, Small Interfering / pharmacology
  • Rats
  • Sodium-Hydrogen Exchangers / antagonists & inhibitors
  • Sodium-Hydrogen Exchangers / chemistry
  • Sodium-Hydrogen Exchangers / metabolism*
  • rho-Associated Kinases

Substances

  • Amides
  • CAV1 protein, human
  • Cav1 protein, rat
  • Caveolin 1
  • Caveolins
  • Enzyme Inhibitors
  • Hyaluronan Receptors
  • Immunoglobulin G
  • Intracellular Signaling Peptides and Proteins
  • Membrane Proteins
  • Pyridines
  • RNA, Small Interfering
  • Sodium-Hydrogen Exchangers
  • flotillins
  • growth factor-activatable Na-H exchanger NHE-1
  • Y 27632
  • Amiloride
  • Hyaluronic Acid
  • Protein Serine-Threonine Kinases
  • rho-Associated Kinases
  • Hyaluronoglucosaminidase
  • Cathepsin B
  • ethylisopropylamiloride