Oscillating Aquaporin Phosphorylation and 14-3-3 Proteins Mediate the Circadian Regulation of Leaf Hydraulics
Résumé
The circadian clock regulates plant tissue hydraulics to synchronize water supply with environmental cycles and thereby optimize growth. A role for aquaporin water channels in these processes was suggested by circadian fluctuations in aquaporin transcript abundance. Here, we show that Arabidopsis rosette hydraulic conductivity (Kros) displays a genuine circadian rhythmicity with a peak around midday. Combined immunological and proteomic approaches revealed that phosphorylation at two C-terminal sites (Ser280, Ser283) of Plasma membrane Intrinsic Protein 2;1 (AtPIP2;1), one of major plasma membrane aquaporins in rosettes, shows circadian oscillations and is correlated with Kros. Transgenic expression of phosphodeficient and phosphomimetic forms of this aquaporin indicated AtPIP2;1 phosphorylation to be necessary but not sufficient for Kros regulation. The supporting role of 14-3-3 proteins, known to interact with and regulate phosphorylated proteins, was investigated. Individual knock-out plants for five 14-3-3 protein isoforms expressed in rosettes lacked circadian activation of Kros. Two of these (GRF4 (14-3-3Phi); GRF10 (14-3-3Epsilon)) showed direct interactions with AtPIP2;1 in the plant and upon co-expression in oocytes were able to activate AtPIP2;1, preferentially when the latter is phosphorylated at its two C-terminal sites. We propose that this regulation mechanism assists in activation of phosphorylated AtPIP2;1 during circadian regulation of Kros.