Colocalization of GP125/CD98 with tropomyosin isoforms at the cell-cell adhesion boundary

Citation
T. Shishido et al., Colocalization of GP125/CD98 with tropomyosin isoforms at the cell-cell adhesion boundary, J BIOCHEM, 127(2), 2000, pp. 253-261
Citations number
60
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
JOURNAL OF BIOCHEMISTRY
ISSN journal
0021-924X → ACNP
Volume
127
Issue
2
Year of publication
2000
Pages
253 - 261
Database
ISI
SICI code
0021-924X(200002)127:2<253:COGWTI>2.0.ZU;2-Q
Abstract
Two monoclonal antibodies designated as 1F6 and 4B10 were obtained on scree ning for reactivities to CD98-associated molecules by sandwich-type enzyme- linked immunosorbent assaying using hybridoma culture supernatants as the s olid phase, cell lysates as an antigen source, and a mixture of biotinylate d antibodies to CD98HC as a detector. Flow cytometric analysis with microsp heres in combination with 1F6, 4B10, and anti-CD98HC also indicated the ass ociation of antibody-defined antigen(s) with CD98, 1F6 and 4B10, stained fi brillate components in fixed and permeated cells but were not reactive with unfixed live cells, suggesting that epitopes reside in the cytoskeleton-as sociated structure in the intracellular region. Two-color immunostaining fo llowed by confocal microscopy revealed the colocalization of the antigen wi th CD98 at the cell-cell adhesion boundary of HeLa cells, 1F6 detected prot eins with relative molecular masses of 33,000 to 43,000 on immunoblotting a nalysis involving cell lysates of human and rat cell lines. Analysis with a purified tropomyosin specimen from rabbit skeletal muscle demonstrated tha t 1F6 and 4B10 recognize tropomyosin, Two-dimensional gel electrophoresis f ollowed by immunoblotting analysis revealed that 1F6 recognizes various tro pomyosin isoforms, These results indicated that CD98 physically associates directly or indirectly with tropomyosin, and that this association is close ly related to the cell-cell interaction.