P2X(7) receptor stimulation upregulates Egr-1 biosynthesis involving a cytosolic Ca(2+) rise, transactivation of the EGF receptor and phosphorylation of ERK and Elk-1.
The P2X(7) receptor is an ATP-gated ionotropic receptor that is permeable for small cations including Ca(2+) ions. Using 293 cells expressing P2X(7) receptors, we show that the P2X(7) receptor-specific ligand 2',3'-O-(4-benzoyl-benzoyl)-ATP (BzATP) induces a signaling cascade leading to the biosynthesis of biologically active Egr-1, a zinc finger transcription factor. BzATP-triggered Egr-1 biosynthesis was attenuated by the mitogen-activated protein kinase kinase inhibitor PD98059, by BAPTA-AM, the acetoxymethylester of the cytosolic Ca(2+) chelator BAPTA, and by an epidermal growth factor (EGF) receptor-specific tyrosine kinase inhibitor (AG1478). These results indicate that phosphorylation and activation of extracellular signal-regulated protein kinase ERK, elevated levels of intracellular Ca(2+) and the transactivation of the EGF receptor are essential for BzATP-induced upregulation of Egr-1. The requirement of Ca(2+) within the signaling cascade was upstream of Raf kinase activation. Lentiviral-mediated expression of MAP kinase phosphatase-1 (MKP-1), a dual-specific phosphatase that dephosphorylates and inactivates ERK in the nucleus, inhibited Egr-1 biosynthesis following BzATP stimulation, indicating that MKP-1 functions as a nuclear shut-off device. Furthermore, the ternary complex factor Elk-1 was phosphorylated and the transcriptional activation potential of Elk-1 was enhanced following P2X(7) receptor stimulation. Expression of a dominant-negative mutant of Elk-1 impaired BzATP-induced upregulation of Egr-1 biosynthesis. Thus, Elk-1 connects the intracellular signaling cascade elicited by activation of P2X(7) receptors with the transcription of the Egr-1 gene.
Stefano
Luisa
L
Department of Medical Biochemistry and Molecular Biology, University of Saarland Medical Center, Homburg, Germany.
Rössler
Oliver G
OG
Griesemer
Désirée
D
Hoth
Markus
M
Thiel
Gerald
G
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D054638:Dual Specificity Phosphatase 1; D051766:Early Growth Response Protein 1 / Q000096:biosynthesis* / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism; D020935:MAP Kinase Signaling System; D008954:Models, Biological; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism*; D058486:Receptors, Purinergic P2X7; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D015533:Transcriptional Activation; D014162:Transfection; D015854:Up-Regulation; D050807:ets-Domain Protein Elk-1 / Q000378:metabolism
D018797:Cell Cycle Proteins; C497325:EGR1 protein, human; C494112:ELK1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; C546468:P2RX7 protein, human; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D011994:Recombinant Proteins; D050807:ets-Domain Protein Elk-1; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D066246:ErbB Receptors; D048049:Extracellular Signal-Regulated MAP Kinases; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; C517019:DUSP1 protein, human; D054638:Dual Specificity Phosphatase 1; D017027:Protein Tyrosine Phosphatases; D002118:Calcium