Palmitoylation modification of Gαo depresses its susceptibility to GAP-43 activation

Interaction between GAP-43 (growth associated protein-43) and Gαo (alpha subunit of Go protein) influences the signal transduction pathways leading to differentiation of neural cells. GAP-43 is known to increase guanine nucleotide exchange by Gαo, which is a major component of neuronal growth cone membranes. However, it is not clear whether GAP-43 stimulation is related to the Gαo palmitoylation or the conversion of Gαo from oligmers to monomers, which was shown to be a necessary regulatory factor in GDP/GTP exchange of Gαo. Here we expressed and purified GAP-43, GST-GAP-43 and Gαo proteins, detected their stimulatory effect on [35S]-GTPγS binding of Gαo. It was found that the EC50 of both GAP-43 and GST-GAP-43 activation were tenfold lower in case of depalmitoylated Gαo than palmitoylated Gαo. Non-denaturing gel electrophoresis and p-PDM cross-linking analysis revealed that addition of GST-GAP-43 induced disassociation of depalmitoylated Gαo from oligomers to monomers, but did not influence the oligomeric state of palmitoylated Gαo, which suggests that palmitoylation is a key regulatory factor in GAP-43 stimulation on Gαo. These results indicated the interaction of GAP-43 and Gαo could accelerate conversion of depalmitoylated Gαo but not palmitoylated Gαo from oligomers to monomers, so as to increase the GTPγS binding activity of Gαo. Results here provide new evidence about how signaling protein palmitoylation is involved in the G-protein-coupled signal transduction cascade, and give a useful clue on the participation of GAP-43 in G-protein cycle by its preferential activation of depalmitoylated Gαo.