Ser6 in the maize actin‐depolymerizing factor, ZmADF3, is phosphorylated by a calcium‐stimulated protein kinase and is essential for the control of functional activity

Abstract

SummaryMaize actin-depolymerizing factor, ZmADF, binds both Gand F-actin and enhances in vitro actin dynamics. Evidence from studies on vertebrate ADF/cofilin supports the view that this class of protein responds to intracellular and extracellular signals and causes actin reorganization. As a test to determine whether such signal-responsive pathways existed in plants, this study addressed the ability of maize ADF to be phosphorylated and the likely effects of such phosphorylation on its capacity to modulate actin dynamics. It is shown that maize ADF3 (ZmADF3) can be phosphorylated by a calcium-stimulated protein kinase present in a 40-70% ammonium sulphate fraction of a plant cell extract. Phosphorylation is shown to be on Ser6, which is only one of nine amino acids that are fully conserved among the ADF/cofilin proteins across distantly related species. In addition, an analogue of phosphorylated ZmADF3 created by mutating Ser6 to Asp6 (zmadf3-4) does not bind G-or F-actin and has little effect on the enhancement of actin dynamics. These results are discussed in context of the previously observed actin reorganization in root hair cells.

Imaging of the Yellow Cameleon 3.6 Indicator Reveals That Elevations in Cytosolic Ca2+ Follow Oscillating Increases in Growth in Root Hairs of Arabidopsis

“…Thus, there is strong evidence supporting a regulatory role for the Ca 21 gradient through imposing spatial control on the site of cell expansion in these tip-growing systems. Indeed, the structures of the apical actin cytoskeleton and its regulatory proteins (villins, gelsolins, and actin-depolymerizing factors; Smertenko et al, 1998;Tominaga et al, 2000;Allwood et al, 2001;Ketelaar et al, 2003;Fan et al, 2004) are thought to be regulated by the tip-high Ca 21 (Yokota et al, 2005). Similarly, there is evidence that annexins (Blackbourn et al, 1992;Clark et al, 1992;Carroll et al, 1998), phosphoinositide metabolism (Preuss et al, 2006), and calmodulin and protein kinases (Moutinho et al, 1998;Yoon et al, 2006) also play roles in sustaining tip growth and are regulated via the cytosolic Ca 21 gradient.…”

14-3-3 λ protein interacts with ADF1 to regulate actin cytoskeleton dynamics in Arabidopsis

“…The activity of ADF proteins is also regulated by phosphorylation in plant [20]. It has been reported that CDPKs phosphorylate ADFs in plant [19,20]. In this study, we identify that 14-3-3  interacts with ADF1 in plant and functions in ADF1 phosphorylation regulation to modulate actin cytoskeleton reorganization.…”

“…In plant, actin interacting proteins (AIP) interact with ADF proteins and enhance their F-actin severing and depolymerizing activity [41][42][43]. The activity of ADF proteins is also regulated by phosphorylation in plant [20]. It has been reported that CDPKs phosphorylate ADFs in plant [19,20].…”

“…The phosphorylation of Ser 6 is required for ADF actin binding and severing activity [18,19]. Phosphorylation is key regulatory mechanism of ADF activity [20,21]. However, these regulation pathways are not well documented in plant.…”

“…However, these regulation pathways are not well documented in plant. In Arabidopsis, ADF1 is possibly phosphorylated by calcium-dependent protein kinase (CDPK) at Ser 6 [19,20].…”