Phospholipase D stimulation by receptor tyrosine kinases mediated by protein kinase C and a Ras/Ral signaling cascade

M Voss, P A Weernink, S Haupenthal, U Möller, R H Cool, B Bauer, J H Camonis, K H Jakobs, M Schmidt

OnderzoeksoutputAcademicpeer review

59 Citaten (Scopus)
11 Downloads (Pure)


Stimulation of phospholipase D (PLD) in HEK-293 cells expressing the M(3) muscarinic receptor by phorbol ester-activated protein kinase C (PKC) apparently involves Ral GTPases. We report here that PKC, but not muscarinic receptor-induced PLD stimulation in these cells, is strongly and specifically reduced by expression of dominant-negative RalA, G26A RalA, as well as dominant-negative Ras, S17N Ras. In contrast, overexpression of the Ras-activated Ral-specific guanine nucleotide exchange factor, Ral-GDS, specifically enhanced PKC-induced PLD stimulation. Moreover, recombinant Ral-GDS potentiated Ral-dependent PKC-induced PLD stimulation in membranes. Epidermal growth factor, platelet-derived growth factor, and insulin, ligands for receptor tyrosine kinases (RTKs) endogenously expressed in HEK-293 cells, apparently use the PKC- and Ras/Ral-dependent pathway for PLD stimulation. First, PLD stimulation by the RTK agonists was prevented by PKC inhibition and PKC down-regulation. Second, expression of dominant-negative RalA and Ras mutants strongly reduced RTK-induced PLD stimulation. Third, overexpression of Ral-GDS largely potentiated PLD stimulation by the RTK agonists. Finally, using the Ral binding domain of the Ral effector RLIP as an activation-specific probe for Ral proteins, it is demonstrated that endogenous RalA is activated by phorbol ester and RTK agonists. Taken together, strong evidence is provided that RTK-induced PLD stimulation in HEK-293 cells is mediated by PKC and a Ras/Ral signaling cascade.

Originele taal-2English
Pagina's (van-tot)34691-34698
Aantal pagina's8
TijdschriftThe Journal of Biological Chemistry
Nummer van het tijdschrift49
StatusPublished - 3-dec-1999
Extern gepubliceerdJa

Citeer dit