A difference of opinion or a matter of perspective

The letter by Drs Chabre, Antonny and Paris raises the question of whether PtdIns(4,5)P2 is an activator or a terminator of small G proteins. To some extent, the answer to this question depends on one's perspective—is the glass half-empty or half-full? What the data obtained for both Arf and Cdc42 (or Rho) show is that the binding of PtdIns(4,5)P2 strongly accelerates the dissociation of GDP from these G proteins. Because GDP release is the rate-limiting step for G-protein activation, we suggested that PtdIns(4,5)P2 might provide an alternative to Dbl-related molecules in catalyzing the activation of Cdc42.Chabre and colleagues argue that PtdIns(4,5)P2, by weakening guanine-nucleotide binding, is a G-protein terminator. They imply that the effects of PtdIns(4,5)P2 are probably due to nonspecific denaturation of the G proteins, as might be expected with `micelles of an anionic detergent like SDS'. However, there are various pieces of evidence presented in Ref. [1xZheng, Y., Glaven, J.A., Wu, J.W., and Cerione, R.A. J. Biol. Chem. 1996; 271: 23815–23819Crossref | PubMed | Scopus (75)See all References][1]that argue against nonspecific denaturation being responsible for the observed effect of PtdIns(4,5)P2 on Cdc42.First, the effects of PtdIns(4,5)P2 were highly specific and were not observed with other negatively charged lipids, including phosphatidylserine and phosphatidylethanolamine. Second, similar effects of PtdIns(4,5)P2 were not seen with many other small G proteins, such as H-Ras, K-Ras, Rap1a and Ran. Third, direct-binding experiments showed that PtdIns(4,5)P2 bound best to the nucleotide-depleted state of Cdc42, in a fashion identical to that of the guanine-nucleotide-exchange factor Dbl. Thus, like Dbl, PtdIns(4,5)P2 binds to GDP-occupied Cdc42, stimulates GDP release and stabilizes the nucleotide-depleted state of this G protein. Finally, and perhaps most importantly, removal of the last seven amino acid residues from the C-terminal region of Cdc42 eliminated PtdIns(4,5)P2 binding and PtdIns(4,5)P2-stimulated GDP dissociation, without affecting Dbl-stimulated GDP association.Taken together, these data indicate that PtdIns(4,5)P2 acts through a specific binding interaction, rather than through nonspecific denaturation of the guanine-nucleotide-binding site of the G protein. Moreover, the fact that binding of PtdIns(4,5)P2 to the C-terminus of Cdc42 influences guanine nucleotide association raises some interesting possibilities regarding the interactions of other regulatory factors with this region of the G protein. In fact, there is already a precedent for binding interactions at the C-terminal regions of Rho proteins having a regulatory influence on guanine nucleotide binding: the ability of the GDI to bind and inhibit GDP dissociation from Cdc42 and other Rho proteins requires the presence of an isoprenoid moiety at their C-terminal cysteine residues.Thus, contrary to the view of Chabre and colleagues, we would argue that the PtdIns(4,5)P2 findings pose a number of interesting questions for future study. These include how other cellular factors (proteins and lipids) might influence the binding of PtdIns(4,5)P2 to Cdc42, whether other related lipids (e.g. PI 3-kinase products) might act more effectively than PtdIns(4,5)P2, whether rapid reductions in PtdIns(4,5)P2 levels result in bona fide GTP–GDP exchange, and whether PtdIns(4,5)P2 or related phosphatidylinositol phosphates anchor Cdc42 to a specific membrane location.