Type 2C Phosphatase 1 of Artemisia annua L. Is a Negative Regulator of ABA Signaling

Figure 3

AaPP2C1 localizes at both cytosol and nucleus and interaction with AaPYL9. (a) Subcellular localization of AaPP2C1 and AaPYL9 in Agrobacterium-infiltrated tobacco leaves. Nucleus was visualized by DAPI staining. Left panel: YFP channel; middle panel: DAPI channel; right panel: merged picture. Treatment with 10 μM ABA for 1 h did not change the subcellular localization of AaPP2C1. (b) The construct of genes fused N- and C-terminal to YFP, respectively (as indicated), were coinfiltrated into tobacco leaves. YFP signal was observed 48 h to 60 h after infiltrated. (c) The BiFC assay between two mutations of AaPP2C1 and AaPYL9. (d) Yeast two-hybrid assay confirming the interaction between AaPP2C1 and AaPYL9. Bait indicates the protein fusion with Gal4 BD domain. Prey indicates the protein fusion with Gal4 AD domain. GBD is empty pGBKT7 vector, GAD is empty pGADT7 vector. (+His, +Ade) indicates Leu-Trp SD medium; (−His, +Ade) indicates Leu-Trp-His SD medium; (−His, −Ade) indicates Leu-Trp-His-Adedeficient SD medium. AaPYL9m1: substitution mutation P89S; AaPYL9m2: substitution mutation H116A. AaPP2C1m1: substitution mutation G199D; AaPP2C1m2: deletion mutation (four amino acid residues from 305 to 308: RGKE).

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