Small GTP-binding protein Ral is involved in cAMP-mediated release of von Willebrand factor from endothelial cells.
OBJECTIVE
von Willebrand factor (vWF) is synthesized by endothelial cells and stored in specialized vesicles called Weibel-Palade bodies (WPBs). Recently, we have shown that the small GTP-binding protein Ral is involved in thrombin-induced exocytosis of WPBs. In addition to Ca2+-elevating secretagogues such as histamine and thrombin, release of WPB is also observed after administration of cAMP-raising substances such as epinephrine and vasopressin. In the present study, we investigated whether Ral is also involved in cAMP-mediated vWF release.


RESULTS
Activation of Ral was observed 15 to 20 minutes after stimulation of endothelial cells with epinephrine, forskolin, or dibutyryl-cAMP. A cell-permeable peptide comprising the carboxy-terminal part of the Ral protein reduced both thrombin-induced and epinephrine-induced vWF secretion supporting a crucial role for Ral in this process. Furthermore, inhibition of protein kinase A by H-89 resulted in a marked reduction of vWF release and greatly diminished levels of GTP-Ral on stimulation with epinephrine. Activation of Ral was independent of the activation of Epac, a cAMP-regulated exchange factor for the small GTPases Rap1 and Rap2.


CONCLUSIONS
These results suggest that protein kinase A-dependent activation of Ral regulates cAMP-mediated exocytosis of WPB in endothelial cells.
Rondaij
Mariska G
MG
Department of Plasma Proteins, Sanquin Research at CLB, Amsterdam, The Netherlands. J.Voorberg@sanquin.nl
Sellink
Erica
E
Gijzen
Karina A
KA
ten Klooster
Jean Paul
JP
Hordijk
Peter L
PL
van Mourik
Jan A
JA
Voorberg
Jan
J
D003994:Bucladesine / Q000494:pharmacology; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000502:physiology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology*; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004837:Epinephrine / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects*; D015696:Gene Products, tat / Q000502:physiology; D006801:Humans; D010446:Peptide Fragments / Q000494:pharmacology; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D015290:Second Messenger Systems / Q000187:drug effects / Q000502:physiology; D013873:Thionucleotides / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D014471:Umbilical Veins; D014667:Vasopressins / Q000494:pharmacology; D020302:Weibel-Palade Bodies / Q000187:drug effects / Q000378:metabolism*; D020725:ral GTP-Binding Proteins / Q000502:physiology*; D014841:von Willebrand Factor / Q000378:metabolism*
D015696:Gene Products, tat; D010446:Peptide Fragments; D013873:Thionucleotides; D014841:von Willebrand Factor; D014667:Vasopressins; D005576:Colforsin; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D003994:Bucladesine; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D013917:Thrombin; D020725:ral GTP-Binding Proteins; D004837:Epinephrine