Crk-associated substrate p130(Cas) interacts with nephrocystin and both proteins localize to cell-cell contacts of polarized epithelial cells

Exp Cell Res. 2000 Apr 10;256(1):168-78. doi: 10.1006/excr.2000.4822.

Abstract

Crk-associated substrate (p130(Cas), Cas) is a docking protein first recognized as having elevated phosphotyrosine content in mammalian cells transformed by v-Src and v-Crk oncoproteins. Subsequent studies have implicated Cas in the control of normal cell behavior through its roles in integrin-mediated signal transduction and organization of the actin cytoskeleton at sites of cell adhesion. In this study, we sought to gain new insight into normal Cas function by identifying previously unrecognized interacting proteins. A yeast two-hybrid screen using the C-terminal region of Cas as a bait identified the Src homology 3 (SH3) domain of the mouse "nephrocystin" protein-orthologous to a human protein whose loss of function leads to the cystic kidney disease familial juvenile nephronophthisis. The putative full-length mouse and partial canine nephrocystin sequences were deduced from cDNA clones. Additional studies using epitope-tagged mouse nephrocystin indicated that nephrocystin and Cas can interact in mammalian cells and revealed that both proteins prominently localize at or near sites of cell-cell contact in polarized Madin-Darby canine kidney epithelial cells. Our findings provide novel insight into the normal cellular activities regulated by both Cas and nephrocystin, and raise the possibility that these proteins have a related function in polarized epithelial cells.

Publication types

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

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Amino Acid Sequence
  • Animals
  • Cell Line
  • Cell Polarity
  • Cell Transformation, Neoplastic
  • Crk-Associated Substrate Protein
  • Dogs
  • Epithelial Cells / cytology*
  • Epithelial Cells / physiology*
  • Epithelial Cells / ultrastructure
  • Genes, src
  • Humans
  • Intercellular Junctions / physiology*
  • Intercellular Junctions / ultrastructure
  • Kidney
  • Membrane Proteins
  • Mice
  • Molecular Sequence Data
  • Oncogene Protein v-crk
  • Phosphoproteins / analysis
  • Phosphoproteins / chemistry
  • Phosphoproteins / metabolism*
  • Proteins / analysis
  • Proteins / chemistry
  • Proteins / metabolism*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Retinoblastoma Protein / metabolism
  • Retinoblastoma-Like Protein p130
  • Retroviridae Proteins, Oncogenic / genetics
  • Retroviridae Proteins, Oncogenic / metabolism*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Transcription, Genetic
  • Transfection
  • src Homology Domains

Substances

  • Adaptor Proteins, Signal Transducing
  • BCAR1 protein, human
  • Bcar1 protein, mouse
  • Crk-Associated Substrate Protein
  • Membrane Proteins
  • NPHP1 protein, human
  • Oncogene Protein v-crk
  • Phosphoproteins
  • Proteins
  • Recombinant Proteins
  • Retinoblastoma Protein
  • Retinoblastoma-Like Protein p130
  • Retroviridae Proteins, Oncogenic