Phosphorylation of maize plasma membrane aquaporins modulates their channel activity and trafficking

(eng) Aquaporins (AQPs) are channel proteins that facilitate the transmembrane movement of water and/or small neutral solutes. Plant AQPs belonging to the plasma membrane intrinsic protein family (PIP) were shown to be regulated by phosphorylation/dephosphorylation mechanisms. The role of phosphorylation of maize ZmPIP2;1 and ZmPIP1;2 on their channel activity, localization and interaction was investigated in Xenopus laevis oocytes. We demonstrated that serines 126 and 203 of ZmPIP2;1 are important for the protein activity and plasma membrane localization. However, serine phosphorylation seemed not involved in the formation of hetero-oligomers with ZmPIP1;2. Wild type and mutated ZmPIP2;1 and ZmPIP1;2 fused to fluorescent tags were transiently expressed in maize mesophyll protoplasts and localized using a confocal microscope. Wild type CFP-ZmPIP2;1 and YFP-ZmPIP1;2 were predominantly localized in the plasma membrane and endoplasmic reticulum, respectively. Mutation of S288 to alanine in CFP-ZmPIP2;1 led to protein internalization, whereas mutations of S203 or S285 enhanced its plasma membrane localization. Interestingly, YFP-ZmPIP1;2 isoforms mutated in S16 and/or S131 were re- located from the endoplasmic reticulum to the plasma membrane. We also demonstrated that phosphorylation of a highly conserved serine located in the cytosolic loop B was important for the water channel activity of ZmPIP2;1 and ZmPIP1;2 in plant cells. Finally, preliminary experiments suggested that phosphorylation status of ZmPIP2s might be altered upon drought or salt stress. Altogether, these data indicate that phosphorylation of PIP proteins play multiple important regulatory roles.