Arabidopsis Phospholipase Dα1 Interacts with the Heterotrimeric G-protein α-Subunit through a Motif Analogous to the DRY Motif in G-protein-coupled Receptors

Phospholipase D (PLD) and heterotrimeric G-protein both play important, diverse roles in cellular regulation and signal transduction. Here we have determined the physical interaction between plant PLD and the only canonical -subunit (G) of the G-protein in Arabidopsis thaliana and the molecular basis for the interaction. PLD1 expressed in either Escherichia coli or Arabidopsis was co-precipitated with G. PLD1 contains a sequence motif analogous to the G-interacting DRY motif normally conserved in G-protein-coupled receptors. Mutation of the central Lys residue PLDK564A of this motif abolished the PLD1-G binding, whereas mutation of the two flanking residues PLDE563A and PLDF565A decreased the binding. Addition of G to PLD1 inhibited PLD1 activity, whereas the PLDK564A mutation that disrupted the G-PLD1 binding abolished the inhibition. GTP relieved the G inhibition of PLD1 activity and also inhibited the binding between PLD1 and G. Meanwhile, the PLD1-G interaction stimulated the intrinsic GTPase activity of G. Therefore, these results have demonstrated the direct binding between G and PLD1, identified the DRY motif on PLD1 as the site for the interaction, and indicated that the interaction modulates reciprocally the activities of PLD1 and G.