PKD is the founding member of a novel protein kinase family that also includes PKD2 and PKD3. PKD has been the focus of most studies up to date, but little is known about the mechanisms that mediate PKD3 activation. Here, we demonstrate that PKD3 immunoprecipitated from COS-7 cells transfected with a constitutively active G alpha q subunit (alpha(q)Q209L) exhibited a marked increase in basal activity. Addition of aluminum fluoride to cells co-transfected with PKD3 and wild type G alpha(q) also induced PKD3 activation. G alpha(q)-mediated PKD3 activation is associated with persistent translocation of PKD3 from both cytosol and nucleus to plasma membrane. Expression of a COOH-terminal fragment of G alpha(q) that acts in a dominant-negative fashion attenuated PKD3 activation in response to bombesin receptor stimulation. Our results indicate that G alpha(q) activation is sufficient to stimulate sustained PKD3 activation and show that the endogenous G alpha q is a major component in the signaling pathway that mediates bombesin-induced PKD3 activation. (c) 2005 Elsevier Inc. All rights reserved.