ATP activates DNA synthesis by acting on P2X receptors in human osteoblast-like MG-63 cells.
In human osteoblast-like MG-63 cells, extracellular ATP increased [(3)H]thymidine incorporation and cell proliferation and synergistically enhanced platelet-derived growth factor- or insulin-like growth factor I-induced [(3)H]thymidine incorporation. ATP-induced [(3)H]thymidine incorporation was mimicked by the nonhydrolyzable ATP analogs adenosine 5'-O-(3-thiotriphosphate) and adenosine 5'-adenylylimidodiphosphate and was inhibited by the P2 purinoceptor antagonist suramin, suggesting involvement of P2 purinoceptors. The P2Y receptor agonist UTP and UDP and a P2Y receptor antagonist reactive blue 2 did not affect [(3)H]thymidine incorporation, whereas the P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2',4-disulfonic acid inhibited ATP-induced [(3)H]thymidine incorporation, suggesting that ATP-induced DNA synthesis was mediated by P2X receptors. RT-PCR analysis revealed that MG-63 cells expressed P2X(4), P2X(5), P2X(6), and P2X(7), but not P2X(1), P2X(2), and P2X(3), receptors. In fura 2-loaded cells, not only ATP, but also UTP, increased intracellular Ca(2+) concentration, and inhibitors for several Ca(2+)-activated protein kinases had no effect on ATP-induced DNA synthesis, suggesting that an increase in intracellular Ca(2+) concentration is not indispensable for ATP-induced DNA synthesis. ATP increased mitogen-activated protein kinase activity in a Ca(2+)-independent manner and synergistically enhanced platelet-derived growth factor- or insulin-like growth factor I-induced kinase activity. Furthermore, the mitogen-activated protein kinase kinase inhibitor PD-98059 totally abolished ATP-induced DNA synthesis. We conclude that ATP increases DNA synthesis and enhances the proliferative effects of growth factors through P2X receptors by activating a mitogen-activated protein kinase pathway.
Department of Orthopedic Surgery, School of Medicine, University of Occupational and Environmental Health, Kitakyushu 807-8555, Japan.
E
E
Nakamura
Y
Y
Uezono
K
K
Narusawa
I
I
Shibuya
Y
Y
Oishi
M
M
Tanaka
N
N
Yanagihara
T
T
Nakamura
F
F
Izumi
D000255:Adenosine Triphosphate / Q000502:physiology*; D004247:DNA / Q000096:biosynthesis* / Q000187:drug effects; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D018048:Receptors, Purinergic P2 / Q000187:drug effects / Q000378:metabolism*; D058485:Receptors, Purinergic P2X5; D014407:Tumor Cells, Cultured
C546465:P2RX5 protein, human; D010982:Platelet-Derived Growth Factor; D018048:Receptors, Purinergic P2; D058485:Receptors, Purinergic P2X5; D007334:Insulin-Like Growth Factor I; D000255:Adenosine Triphosphate; D004247:DNA; D020928:Mitogen-Activated Protein Kinases