Direct binding of neutral endopeptidase 24.11 to ezrin/radixin/moesin (ERM) proteins competes with the interaction of CD44 with ERM proteins.
Neutral endopeptidase 24.11 (NEP) is a cell surface peptidase expressed by numerous tissues including prostatic epithelial cells. We reported that NEP inhibits prostate cancer cell proliferation and cell migration by enzymatic inactivation of neuropeptide substrates and through protein-protein interaction independent of catalytic function. The cytoplasmic domain of NEP contains a positively charged amino acid cluster, previously identified as a binding site for ezrin/radixin/moesin (ERM) proteins. We report here that NEP co-immunoprecipitates with ERM proteins in NEP-expressing LNCaP prostate cancer cells and MeWo melanoma cells. Co-immunoprecipitation showed that ERM proteins associate with wild-type NEP protein but not NEP protein containing a truncated cytoplasmic domain or point mutations replacing the positively charged amino acid cluster. In vitro binding assays showed that NEP binds directly to recombinant N terminus fragments of ERM proteins at the positively charged amino acid cluster within the NEP cytoplasmic domain. Binding of ERM proteins to NEP results in decreased binding of ERM proteins to the hyaluronan receptor CD44, a main binding partner of ERM proteins. Moreover, cells expressing wild-type NEP demonstrate decreased adhesion to hyaluronic acid and cell migration. These data suggest that NEP can affect cell adhesion and migration through direct binding to ERM proteins.
Iwase
Akira
A
Urologic Oncology Research Laboratory, Department of Urology, Weill Medical College of Cornell University, New York, New York 10021, USA.
Shen
Ruoqian
R
Navarro
Daniel
D
Nanus
David M
DM
D001483:Base Sequence; D001667:Binding, Competitive; D001798:Blood Proteins / Q000378:metabolism*; D002460:Cell Line; D003593:Cytoplasm / Q000201:enzymology; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D017931:DNA Primers; D018960:Hyaluronan Receptors / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008840:Microfilament Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D015260:Neprilysin / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D000099149:Ezrin
D001798:Blood Proteins; D003598:Cytoskeletal Proteins; D017931:DNA Primers; D018960:Hyaluronan Receptors; D008565:Membrane Proteins; D008840:Microfilament Proteins; D015260:Neprilysin; D010750:Phosphoproteins; D000099149:Ezrin; C071058:moesin; C071776:radixin