Retinoids increase cell-cell adhesion strength, beta-catenin protein stability, and localization to the cell membrane in a breast cancer cell line: a role for serine kinase activity.
In this study we show that a breast cancer cell line (SKBR3) that expresses no E-cadherin and very low levels of beta-catenin protein and exhibits a poorly adhesive phenotype in Matrigel responds to retinoic acid (RA) by a marked increase in epithelial differentiation. Specifically, treatment of cells with all-trans-RA, 9-cis-RA, or a RA receptor alpha-specific ligand resulted in a large increase in cell-cell adhesive strength and stimulated the formation of fused cell aggregates in Matrigel. A retinoid X receptor-specific ligand was ineffective. Exposure of cells to 9-cis-RA for as little as 4 h was sufficient to maintain the adhesive phenotype for at least 4 days. The effects of 9-cis-RA required protein and RNA synthesis, but were not mediated by factors secreted by stimulated cells or by direct cell contact and did not require serum. These 9-cis-RA-induced morphological effects were completely reversed by growing cells in 50 microM Ca2+, suggesting a mechanism involving a 9-cis-RA-induced increase in Ca(2+)-dependent adhesion. Consistent with this, beta-catenin protein levels were markedly elevated in the 9-cis-RA-treated cells, and beta-catenin became localized to a Triton-insoluble pool at regions of cell-cell contact. No change could be detected in beta-catenin steady state messenger RNA levels, but 9-cis-RA did increase beta-catenin protein stability. Treatment of cells with low calcium medium did not prevent the 9-cis-RA-induced increase in total beta-catenin protein, but did prevent its movement to a Triton-insoluble pool at the cell membrane. Among several kinase inhibitors, only the broad spectrum kinase inhibitor staurosporine and the protein kinase C inhibitor bisindoylmaleimide reversed the morphological changes induced by 9-cis-RA. Like treatment with low calcium medium, these inhibitors did not prevent the 9-cis-RA-induced increase in total beta-catenin protein levels, but completely prevented the movement of beta-catenin to the cell membrane. These results point to a role for beta-catenin and serine kinase activity in mediating the action of 9-cis-RA in epithelial differentiation.
Byers
S
S
Department of Cell Biology, Georgetown University Medical Center, Washington D.C.20007, USA.
Pishvaian
M
M
Crockett
C
C
Peer
C
C
Tozeren
A
A
Sporn
M
M
Anzano
M
M
Lechleider
R
R
D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D015820:Cadherins / Q000378:metabolism; D002118:Calcium / Q000008:administration & dosage / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects; D002454:Cell Differentiation; D002462:Cell Membrane / Q000378:metabolism; D003470:Culture Media; D003598:Cytoskeletal Proteins / Q000187:drug effects / Q000378:metabolism*; D004355:Drug Stability; D004848:Epithelium / Q000187:drug effects / Q000473:pathology; D005260:Female; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D019311:Staurosporine / Q000494:pharmacology; D014018:Tissue Distribution; D015534:Trans-Activators; D014212:Tretinoin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000187:drug effects; D051176:beta Catenin
C495270:CTNNB1 protein, human; D015820:Cadherins; D003470:Culture Media; D003598:Cytoskeletal Proteins; D007211:Indoles; D008301:Maleimides; D012333:RNA, Messenger; D015534:Trans-Activators; D051176:beta Catenin; D014212:Tretinoin; D017346:Protein Serine-Threonine Kinases; D019311:Staurosporine; C088060:bisindolylmaleimide; D002118:Calcium