A collagen peptide motif activates tyrosine kinase-dependent calcium signalling pathways in human osteoblast-like cells.
A collagen peptide motif (DGEA) which is a putative alpha 2 beta 1 integrin binding site was examined for its ability to activate Ca2+ signalling pathways in the human osteoblast-like cell line SaOS-2. We show that these cells express both alpha 2 beta 1 integrin subunits (by immunocytochemistry) and that an anti-beta 1 monoclonal antibody (DF5) mobilizes Ca2+ in these cells. DGEA elevated intracellular Ca2+ in fura-2-loaded cells, in a concentration- and sequence-dependent fashion, with an EC50 of 250 microM. The tyrosine kinase inhibitor herbimycin A reduced the number of cells responding to DGEA and to transforming growth factor alpha. Thrombin also stimulated a rise in intracellular Ca2+, but the number of cells responding was not reduced by herbimycin A. The DGEA response was dependent on extracellular Ca2+, but was not due to Ca2+ influx, since it was blocked by thapsigargin and not by lanthanum. Using three different anti-alpha 2 monoclonal antibodies, we were unable to show that the DGEA-induced Ca2+ signal was mediated by the alpha 2 beta 1 integrin. In summary, the DGEA collagen motif does appear to activate receptor-mediated Ca2+ signalling events in SaOS-2 cells, in a divalent cation-dependent manner, but we were unable to demonstrate a role for alpha 2 beta 1 integrin in this response.
McCann
T J
TJ
Department of Orthodontics and Pediatric Dentistry, United Medical School, London, UK.
Mason
W T
WT
Meikle
M C
MC
McDonald
F
F
D016227:Benzoquinones; D002118:Calcium / Q000502:physiology*; D002461:Cell Line, Transformed; D003094:Collagen / Q000502:physiology*; D006801:Humans; D016023:Integrins / Q000096:biosynthesis; D047029:Lactams, Macrocyclic; D010006:Osteoblasts / Q000201:enzymology* / Q000378:metabolism* / Q000502:physiology; D012516:Osteosarcoma; D010455:Peptides / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011809:Quinones / Q000494:pharmacology; D038981:Receptors, Collagen; D017828:Rifabutin / Q000031:analogs & derivatives; D015398:Signal Transduction / Q000187:drug effects; D014407:Tumor Cells, Cultured
D016227:Benzoquinones; D016023:Integrins; D047029:Lactams, Macrocyclic; D010455:Peptides; D011809:Quinones; D038981:Receptors, Collagen; D017828:Rifabutin; C020754:herbimycin; D003094:Collagen; D011505:Protein-Tyrosine Kinases; D002118:Calcium