Identification of CD9 extracellular domains important in regulation of CHO cell adhesion to fibronectin and fibronectin pericellular matrix assembly

George Cook, Celia M. Longhurst, Svetozar Grgurevich, Shila Cholera, Joseph T. Crossno, Lisa K. Jennings

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

CD9, a 24-kDa member of the tetraspanin family, influences cellular growth and development, activation, adhesion, and motility. Our investigation focuses on the hypothesis that the CD9 second extracellular loop (EC2) is important in modulating cell adhesive events. Using a Chinese hamster ovary (CHO) cell expression system, we previously reported that CD9 expression inhibited cell adhesion to fibronectin and fibronectin matrix assembly. For the first time, a functional epitope on CD9 EC2 that regulates these processes is described. Binding of mAb7, an EC2-specific anti-CD9 monoclonal antibody, reversed the CD9 inhibitory activity on CHO cell adhesion and fibronectin matrix assembly. This reversal of cell phenotype also was observed in CHO cells expressing CD9 EC2 truncations. Furthermore, our data showed that the EC2 sequence 173LETFTVKSCPDAIKEVFDNK192 was largely responsible for the CD9-mediated CHO cell phenotype. Two peptides, 135K-V172 (peptide 5b) and 168P-I185 (peptide 6a), selectively blocked mAb7 binding to soluble CD9 and to CD9 on intact cells. These active peptides reversed the influence of CD9 expression on CHO cell adhesion to fibronectin. In addition, confocal microscopy revealed that CD9 colocalized with the integrin α5β1 and cytoskel etal F-actin in punctate clusters on the cell surface, particularly at the cell margins. Immunoprecipitation studies confirmed CD9 association with β1 integrin. The cellular distribution and colocalization of focal adhesion kinase and a-actinin with cytoskeletal actin was also influenced by CD9 expression. Thus, CD9 may exhibit its effect by modulating the composition of adhesive complexes important in facilitating cell adhesion and matrix assembly.

Original languageEnglish (US)
Pages (from-to)4502-4511
Number of pages10
JournalBlood
Volume100
Issue number13
DOIs
StatePublished - Dec 15 2002

Fingerprint

Cell adhesion
Cricetulus
Fibronectins
Cell Adhesion
Ovary
Peptides
Integrins
Actins
Adhesives
Cells
Cell-Matrix Junctions
Actinin
Focal Adhesion Protein-Tyrosine Kinases
Confocal microscopy
Epitopes
Adhesion
Chemical activation
Monoclonal Antibodies
Association reactions
Phenotype

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

Cite this

Identification of CD9 extracellular domains important in regulation of CHO cell adhesion to fibronectin and fibronectin pericellular matrix assembly. / Cook, George; Longhurst, Celia M.; Grgurevich, Svetozar; Cholera, Shila; Crossno, Joseph T.; Jennings, Lisa K.

In: Blood, Vol. 100, No. 13, 15.12.2002, p. 4502-4511.

Research output: Contribution to journalArticle

Cook, George ; Longhurst, Celia M. ; Grgurevich, Svetozar ; Cholera, Shila ; Crossno, Joseph T. ; Jennings, Lisa K. / Identification of CD9 extracellular domains important in regulation of CHO cell adhesion to fibronectin and fibronectin pericellular matrix assembly. In: Blood. 2002 ; Vol. 100, No. 13. pp. 4502-4511.
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AU - Crossno, Joseph T.

AU - Jennings, Lisa K.

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