Investigation of the effect of the farnesyl protein transferase inhibitor R115777 on isoprenylation and intracellular signalling by the prostacyclin receptor.
The human (h) and mouse (m) prostacyclin receptors (IPs) undergo isoprenylation through attachment of a C-15 farnesyl moiety within their conserved carboxyl terminal -CSLC sequences. Herein, the effects of a novel farnesyl transferase inhibitor R115777 on signalling by the hIP and mIP, overexpressed in human embryonic kidney 293 cells, and by the hIP endogenously expressed in human erythroleukaemia cells were investigated. R115777 significantly impaired IP-mediated cyclic AMP generation (IC(50) 0.37-0.60 nm) and intracellular calcium ([Ca(2+)](i)) mobilization (IC(50) 37-65 nm), but had no effect on signalling by the control nonisoprenylated beta(2) adrenergic receptor or the alpha or beta isoforms of the human thromboxane A(2) receptor (TP). Additionally, R115777 significantly reduced IP-mediated cross-desensitization of signalling by the TP alpha, but not by the TP beta, isoform of the human TP and impaired the farnesylation-dependent processing of the chaperone HDJ-2 protein (IC(50) 4.5 nm). Furthermore, R115777 fully impaired isoprenylation of both the Ha-Ras(WT) and Ha-Ras(CSLC) in vitro and in whole cells confirming that, unlike N-Ras and Ki-Ras, the -CSLC motif associated with the IP cannot support alternative geranylgeranylation in the presence of R115777 and does not act as a substrate for geranylgeranyl transferase 1 in vitro or in whole cells. In conclusion, these data confirm that R115777 potently impairs IP isoprenylation and signalling, and suggest that clinically it may not only target Ras proteins but may also disrupt IP isoprenylation, events which could impact on physiologic processes in which prostacyclin and its receptor are implicated.
O'Meara
Sarah J
SJ
Department of Biochemistry, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
Kinsella
B Therese
BT
D019883:Alkyl and Aryl Transferases / Q000037:antagonists & inhibitors* / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D016285:Iloprost / Q000494:pharmacology; D007494:Ireland; D051379:Mice; D018832:Molecular Chaperones / Q000187:drug effects / Q000378:metabolism; D063065:Organophosphonates; D017368:Protein Prenylation / Q000187:drug effects; D015363:Quinolones / Q000737:chemistry / Q000494:pharmacology*; D020239:Receptor Cross-Talk / Q000187:drug effects / Q000502:physiology; D011982:Receptors, Prostaglandin / Q000145:classification / Q000187:drug effects / Q000235:genetics; D013997:Time Factors; D014162:Transfection / Q000379:methods; D014166:Transferases / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology*; D014316:Tritium
C436083:2-(2-oxo-2-((3,7,11-trimethyl-2,6,10-dodecatrienyl)oxy)aminoethyl)phosphonic acid, (2,2-dimethyl-1-oxopropoxy)methyl ester sodium; D004791:Enzyme Inhibitors; D018832:Molecular Chaperones; D063065:Organophosphonates; D015363:Quinolones; D011982:Receptors, Prostaglandin; D014316:Tritium; D000242:Cyclic AMP; D014166:Transferases; D019883:Alkyl and Aryl Transferases; C064911:p21(ras) farnesyl-protein transferase; D016285:Iloprost; C402769:tipifarnib; D002118:Calcium