Differences in intracellular calcium signaling after activation of the thrombin receptor by thrombin and agonist peptide in osteoblast-like cells.
Thrombin and the thrombin receptor agonist peptide (TRAP) caused a rise in intracellular calcium concentration ([Ca2+]i) in the human osteoblast-like cell line Saos-2. Striking differences in the [Ca2+]i signals elicited by these agonists were revealed. In cell populations, thrombin induced a transient increase in [Ca2+]i while TRAP caused a biphasic [Ca2+]i response consisting of an initial peak and a sustained plateau phase. In individual cells, thrombin mainly caused a single [Ca2+]i transient while TRAP induced repetitive [Ca2+]i spikes. Neither tyrosine phosphorylation, cAMP-dependent phosphorylation, nor pertussis toxin-sensitive G proteins appeared to be involved in thrombin receptor [Ca2+]i signaling in this cell line. However, the sustained [Ca2+]i response caused by TRAP was converted into a transient, thrombin-like response by pretreatment with serine/threonine phosphatase inhibitors. Pretreatment with the phorbol ester phorbol 12-myristate 13-acetate (PMA) abrogated thrombin receptor [Ca2+]i signaling, and TRAP-induced Ca2+ entry was inhibited by the acute treatment with PMA. In contrast, Ca2+ entry stimulated by thapsigargin was not sensitive to agents affecting serine/threonine phosphorylation. The observation that thrombin and TRAP, despite being agonists for a common receptor, induce dissimilar [Ca2+]i responses indicates that binding of TRAP alone is insufficient to fully regulate the thrombin receptor in Saos-2 cells.
Jenkins
A L
AL
Department of Hematology, University of Cambridge, United Kingdom.
Bootman
M D
MD
Berridge
M J
MJ
Stone
S R
SR
D000595:Amino Acid Sequence; D002118:Calcium / Q000502:physiology*; D000242:Cyclic AMP / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D010006:Osteoblasts / Q000502:physiology*; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D010766:Phosphorylation; D019000:Phosphotyrosine; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D018179:Receptors, Thrombin / Q000378:metabolism*; D015398:Signal Transduction; D013729:Terpenes / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019284:Thapsigargin; D013917:Thrombin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000031:analogs & derivatives / Q000502:physiology
D010455:Peptides; D018179:Receptors, Thrombin; D013729:Terpenes; D019000:Phosphotyrosine; D014443:Tyrosine; D019284:Thapsigargin; D000242:Cyclic AMP; D017346:Protein Serine-Threonine Kinases; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium