Molecular cloning and characterization of DdCAD-1, a Ca2+-dependent cell-cell adhesion molecule, in Dictyostelium discoideum

Estella F.S. Wong, Simuran K. Brar, Hiromi Sesaki, Chunzhong Yang, Chi Hung Siu

Research output: Contribution to journalArticlepeer-review

Abstract

Dictyostelium discoideum expresses EDTA-sensitive cell-cell adhesion sites soon after the initiation of development, and a Ca2+-binding protein of M(r) 24,000 (designated DdCAD-1) has been implicated in this type of adhesiveness. We have previously purified DdCAD-1 to homogeneity and characterized its cell binding activity (Brat, S. K., and Siu, C.-H. (1992) J. Biol. Chem. 268, 24902-24909). In this report, we describe the cloning of DdCAD-1 cDNAs. DNA sequencing revealed a single open reading frame coding for a polypeptide containing 213 amine acids. The identity of the cDNA was confirmed by amine acid sequences of two cyanogen bromide peptides. The deduced amine acid sequence of DdCAD-1 exhibits a relatively high degree of sequence similarity with members of the cadherin family and protein S of Myxococcus xanthus. Unlike the other cadherins, the carboxyl-terminal region of DdCAD-1 contains a Ca2+-binding motif. Although analyses of the sequence suggest that the polypeptide lacks a signal peptide sequence and a transmembrane domain, immunofluorescence microscopy demonstrates the association of DdCAD-1 with the ecto-surface of the plasma membrane. To investigate the structure/function relationships of DdCAD-1, glutathione S- transferase fusion proteins containing different DdCAD-1 fragments were expressed and assayed for their 45Ca2+ and cell binding activities. These studies revealed that the cell binding activity is dependent on the amino- terminal segment and not the carboxyl-terminal Ca2+-binding domain and showed additional Ca2+-binding site(s) within the amino-terminal segment.

Original languageEnglish (US)
Pages (from-to)16399-16408
Number of pages10
JournalJournal of Biological Chemistry
Volume271
Issue number27
DOIs
StatePublished - 1996
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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