A phospholipase D and protein kinase C inhibitor blocks the spreading of murine mammary adenocarcinoma cells altering f-actin and beta1-integrin point contact distribution.
Spreading is a critical process involved in motility and growth of tumor cells during the metastatic cascade. Focal adhesion kinase, src-proteins and PKC have been reported to participate in the regulation of cytoskeleton organization in both normal and transformed cells during spreading. The role of other signaling enzymes such as PLD and PAP has not been studied during spreading in tumor cells. We now show that the spreading of murine mammary adenocarcinoma LM3 cells was significantly reduced by n-butanol, a PLD and PKC inhibitor, with a maximal inhibition of 54% (p < 0.001) in both the presence and absence of serum, as measured by phase-contrast microscopy. PMA only stimulated cell spreading over the control in the absence of serum and n-butanol inhibition was completely reversed by PMA treatment in both conditions. PA, the product of PLD activity, stimulated LM3 cell spreading and the same effect was observed with staurosporine. Spreading was enhanced when cells were seeded on collagen-IV- or fibronectin-coated surfaces and n-butanol could inhibit both integrin-derived signals. Cell spreading inhibition correlated with the absence of f-actin bundles and fewer beta1-integrin point contacts as determined by double immunofluorescence microscopy. In addition, n-butanol inhibited the proliferation of LM3 cells in the presence of serum (p < 0.01). These results suggest that beta1-integrin and f-actin/point contact assembly, involved in spreading and proliferation, require the participation of PLD-PKC regulatory pathways in LM3 cells.
Aguirre Ghiso
J A
JA
Research Area, Institute of Oncology Angel H. Roffo University of Buenos Aires, Argentina. postmast@butnol.fmed.uba.ar
Farías
E F
EF
Alonso
D F
DF
Arregui
C
C
Bal de Kier Joffé
E
E
D020001:1-Butanol; D000199:Actins / Q000032:analysis*; D000230:Adenocarcinoma / Q000473:pathology*; D000818:Animals; D000440:Butanols / Q000494:pharmacology; D002448:Cell Adhesion; D002455:Cell Division / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein; D019012:Integrin beta1 / Q000032:analysis*; D007529:Isoflavones / Q000494:pharmacology; D008325:Mammary Neoplasms, Experimental / Q000473:pathology*; D051379:Mice; D008856:Microscopy, Fluorescence; D010712:Phosphatidic Acids / Q000494:pharmacology; D010739:Phospholipase D / Q000037:antagonists & inhibitors*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000199:Actins; D000440:Butanols; D004791:Enzyme Inhibitors; D019012:Integrin beta1; D007529:Isoflavones; D010712:Phosphatidic Acids; D020001:1-Butanol; D019833:Genistein; D011493:Protein Kinase C; D010739:Phospholipase D; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate