NEW PLAYERS IN PLANT AQUAPORIN TRAFFICKING TO THE PLASMA MEMBRANE​

Plasma membrane intrinsic proteins (PIPs) are aquaporins facilitating the passive diffusion of water and/or small neutral solutes across biological membranes. These proteins assemble as tetramers, in which each subunit forms a functional channel. Each monomer is constituted of six transmembrane domains (TM1–TM6), which are linked by five loops (A–E). PIPs cluster into two phylogenetic groups (PIP1 and PIP2) that physically interact to modify their trafficking to the plasma membrane. When transiently expressed in single maize cells, ZmPIP1s are retained in the endoplasmic reticulum (ER), whereas ZmPIP2s are targeted to the PM. Recently, we discovered a new ER export motif (LxxxA) in the TM3 of ZmPIP2;5 (Chevalier et al., 2014, Plant Physiol, 166, 125-138). The mutated form of ZmPIP2;5 (mYFP:ZmPIP2;5L127F/A131M) is retained in intracellular structures colocalizing with markers corresponding to the ER. We are currently working to understand how this TM motif controls the ER exit of PIPs. The contribution of lipids on PIPs trafficking is investigated using lipid biosynthesis inhibitors and membrane lipid strips. We are also setting a proteomic approach to screen for PIP interacting proteins in a cellular environment. This approach, named BioID for proximity dependent biotin identification, was successfully used on animals systems and we are implementing it in plant cell cultures. BioID is based on the fusion of a promiscuous biotin protein ligase to the protein of interest. Consequently, in the presence of biotin, the proteins that are interacting with the fusion protein are biotinylated and could be easily purified and identified by mass spectrometry. The strategy and the first results will be described