CD44v(3,8-10) is involved in cytoskeleton-mediated tumor cell migration and matrix metalloproteinase (MMP-9) association in metastatic breast cancer cells.

Abstract

In the present study, we have employed a unique breast cancer cell line (Met-1, which was derived from a high metastatic potential tumor in transgenic mice expressing polyomavirus middle T oncogene) to study the role of CD44 variant isoform(s) in the regulation of metastatic breast tumor cell behavior. The results of reverse transcriptase-polymerase chain reaction, Southern blot, nucleotide sequencing, immunoprecipitation, and immunoblot analyses indicated that these cells express a major CD44 isoform (molecular weight approximately 260 kDa) containing a v3,8-10 exon insertion (designated as CD44v3,8-10). In addition, we have determined that CD44v3,8-10 binds specifically to the cytoskeletal proteins such as ankyrin. Biochemical analyses, using competition binding assays and a synthetic peptide identical to NGGNGTVEDRKPSEL (a sequence located between aa480 and aa494 of CD44v3,8-10) indicate that this 15-amino acid peptide binds specifically to the cytoskeletal protein ankyrin (but not to fodrin or spectrin). This peptide competes effectively for ankyrin binding to CD44v3,8-10. Therefore, we believe that the sequence 480NGGNGTVEDRKPSE494L, located at the cytoplasmic domain of CD44v3,8-10, is required for the ankyrin binding. We have also detected that CD44v3,8-10-containing Met-1 cells are capable of forming membrane spikes or "invadopodia" structures and undergo active migration processes. Treatments of Met-1 cells with certain agents including anti-CD44v3 antibody, cytochalasin D (a microfilament inhibitor), and W-7 (a calmodulin antagonist), but not colchicine (a microtubule disrupting agent) effectively inhibit "invadopodia" formation and subsequent tumor cell migration. Further analyses using zymography assays and double immunofluorescence staining indicated that CD44v3,8-10 is closely associated with the active form of matrix metalloproteinase, MMP-9, in a complex within "invadopodia" structures. These findings suggest that CD44v3,8-10 plays an important role in linking ankyrin to the membrane-associated actomyosin contractile system required for "invadopodia" formation (coupled with matrix degradation activities) and tumor cell migration during breast cancer progression.

• Authors: L Y Bourguignon, Z Gunja-Smith, N Iida, H B Zhu, L J Young, W J Muller, R D Cardiff
• Journal: Journal of cellular physiology (J Cell Physiol) Vol. 176 Issue 1 Pg. 206-15 (Jul 1998) ISSN: 0021-9541 [Print] United States
• PMID: 9618160 (Publication Type: Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Ankyrins
• Hyaluronan Receptors
• Peptide Fragments
• Sulfonamides
• Cytochalasin D
• W 7
• Collagenases
• Matrix Metalloproteinase 9

Topics

• Amino Acid Sequence
• Animals
• Ankyrins (metabolism)
• Binding, Competitive
• Cell Movement (drug effects, physiology)
• Collagenases (metabolism)
• Cytochalasin D (pharmacology)
• Cytoskeleton (physiology)
• Gene Expression Regulation, Neoplastic (genetics)
• Hyaluronan Receptors (genetics, immunology)
• Immunohistochemistry
• Matrix Metalloproteinase 9
• Mice
• Molecular Sequence Data
• Neoplasm Metastasis (physiopathology)
• Neoplasms, Experimental (metabolism)
• Peptide Fragments (metabolism)
• Protein Binding (physiology)
• Sulfonamides (pharmacology)
• Tumor Cells, Cultured