Structural and functional insights into the regulation of Arabidopsis AGC VIIIa kinases.
The AGCVIIIa kinases of Arabidopsis are members of the eukaryotic PKA, PKG, and PKC group of regulatory kinases. One AGCVIIIa kinase, PINOID (PID), plays a fundamental role in the asymmetrical localization of membrane proteins during polar auxin transport. The remaining 16 AGCVIIIa genes have not been associated with single mutant phenotypes, suggesting that the corresponding kinases function redundantly. Consistent with this idea, we find that the genes encoding the Arabidopsis AGCVIIIa kinases have spatially distinct, but overlapping, expression domains. Here we show that the majority of Arabidopsis AGCVIIIa kinases are substrates for the 3-phosphoinositide-dependent kinase 1 (PDK1) and that trans-phosphorylation by PDK1 correlates with activation of substrate AGCVIIIa kinases. Mutational analysis of two conserved regulatory domains was used to demonstrate that sequences located outside of the C-terminal PDK1 interaction (PIF) domain and the activation loop are required for functional interactions between PDK1 and its substrates. A subset of GFP-tagged AGCVIIIa kinases expressed in Saccharomyces cerevisiae and tobacco BY-2 cells were preferentially localized to the cytoplasm (AGC1-7), nucleus (WAG1 and KIPK), and the cell periphery (PID). We present evidence that PID insertion domain sequences are sufficient to direct the observed peripheral localization. We find that PID specifically but non-selectively binds to phosphoinositides and phosphatidic acid, suggesting that PID might directly interact with the plasma membrane through protein-lipid interactions. The initial characterization of the AGCVIIIa kinases presented here provides a framework for elucidating the physiological roles of these kinases in planta.
Zegzouti
Hicham
H
Department of Molecular, Cell, and Developmental Biology, David Geffen School of Medicine, University of California, Los Angeles 90095, USA.
Li
Wei
W
Lorenz
Todd C
TC
Xie
Mingtang
M
Payne
C Thomas
CT
Smith
Kelly
K
Glenny
Scott
S
Payne
Gregory S
GS
Christensen
Sioux K
SK
D064413:3-Phosphoinositide-Dependent Protein Kinases; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology*; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D021381:Protein Transport
D011494:Protein Kinases; D064413:3-Phosphoinositide-Dependent Protein Kinases; D017346:Protein Serine-Threonine Kinases