Interaction of maize actin-depolymerising factor with actin and phosphoinositides and its inhibition of plant phospholipase C.

We have previously reported the isolation of three Zea mays genes that enco de actin-depolymerising factors/ cofilins, a family of low molecular weight actin regulating proteins. In the present study, we have characterised one of these proteins, ZmADF3. We report that ZmADF3 binds G-actin with a 1:1 stoichiometry, and that the interaction with F-actin is pH-sensitive. ZmADF 3 co-sediments mainly with F-actin at pH 6.0 and mainly with G-actin at pH 9.0. This response is more similar to that of vertebrate cofilin and ADF th an to that of Acanthamoeba actophorin which, although more similar in prima ry sequence to ZmADF3, is not pH sensitive. However, ZmADF3 requires a more basic environment to depolymerise actin relative to either vertebrate ADF or cofilin. Filaments decorated with ZmADF3 at low pH are very rapidly depo lymerised upon raising the pH, which is consistent with a severing mechanis m for the disruption of actin filaments. Also, we demonstrate that ZmADF3 b inds specific polyphosphatidylinositol lipids, especially phosphatidylinosi tol 4,5-bisphosphate (PIP2), and we show that this binding inhibits the act in-depolymerising function of ZmADF3. Moreover, we show that a consequence of ZmADF3 binding PIP2 is the inhibition of the activity of polyphosphatidy linositol specific plant phospholipase C, indicating the possibility of rec iprocal modulation of this major signalling pathway and the actin cytoskele ton.