Maize [Zea mays] plasma membrane aquaporins belonging to the PIP1 and PIP2 subgroups are in vivo phosphorylated

• Published in: Plant and cell physiology 2008-09, Vol.49 (9), p.1364-1377
• Main Authors: Wilder, V.V.(Universite catholique de Louvain (Belgium)), Miecielica, U, Degand, H, Derua, R, Waelkens, E, Chaumont, F
• Format: Article
• Language: English
• Subjects: Animals; Aquaporins; Aquaporins - genetics; Aquaporins - metabolism; BROTES; Cell Membrane - metabolism; Cloning, Molecular; Cyclic AMP-Dependent Protein Kinases; DNA, Complementary - genetics; DNA, Plant - genetics; EXPERIMENTACION IN VITRO; EXPERIMENTACION IN VIVO; EXPERIMENTATION IN VITRO; EXPERIMENTATION IN VIVO; FOSFORILACION; GENE; GENES; Genes, Plant; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; IN VITRO EXPERIMENTATION; IN VIVO EXPERIMENTATION; Maize; Mass Spectrometry; Oocytes - metabolism; Osmosis; OVA; OVULE; OVULO; PEPTIDE; PEPTIDES; PEPTIDOS; PHOSPHORYLATION; Plant Proteins - genetics; Plant Proteins - metabolism; Plant Shoots - genetics; Plant Shoots - metabolism; POUSSE; Protein Kinases - metabolism; PROTEINAS; PROTEINE; PROTEINS; Serine - metabolism; SHOOTS; TRANSFERASAS; TRANSFERASE; TRANSFERASES; Xenopus laevis - genetics; Xenopus laevis - metabolism; Xenopus laevis oocytes; ZEA MAYS; Zea mays - genetics; Zea mays - metabolism
• ISSN: 0032-0781; 1471-9053
• DOI: 10.1093/pcp/pcn112

Aquaporins are channel proteins that facilitate transmembrane water movement. In this study, we showed that plasma membrane intrinsic proteins (PIPs) from maize shoots are in vitro and in vivo phosphorylated on serine residues by a calcium-dependent kinase associated with the membrane fraction. Mass spectrometry identified phosphorylated peptides corresponding to the C-terminal region of (i) ZmPIP2;1, ZmPIP2;2 and/or ZmPIP2;7; (ii) ZmPIP2;3 and/or ZmPIP2;4; (iii) ZmPIP2;6; together with (iv) a phosphorylated peptide located in the N-terminal region of ZmPIP1;1, ZmPIP1;2, ZmPIP1;3 and/or ZmPIP1;4. The role of phosphorylation in the water channel activity of wild-type and mutant ZmPIP2;1 was studied in Xenopus laevis oocytes. Activation of endogenous protein kinase A increased the osmotic water permeability coefficient of ZmPIP2;1-expressing oocytes, suggesting that phosphorylation activates its channel activity. Mutation of S126 or S203, putative phosphorylated serine residues conserved in all plant PIPs, to alanine decreased ZmPIP2;1 activity by 30-50%, without affecting its targeting to the plasma membrane. Mutation of S285, which is phosphorylated in planta, to alanine or glutamate did not affect the water channel activity. These results indicate that, in oocytes, S126 and S203 play an important role in ZmPIP2;1 activity and that phosphorylation of S285 is not required for its activity.