Aquaporins

Aquaporins (AQPs) are channels facilitating the movement of water and small solutes across cellular membranes. Plants appear to express a surprisingly high number of AQP homologues. On the basis of sequence comparison, plant AQPs are classified into four subfamilies, the plasma membrane intrinsic proteins (PIPs), the tonoplast intrinsic proteins (TIPs), the NOD26-like intrinsic proteins (NIPs) and a small group named the small basic intrinsic proteins (SIPs). AQPs are thought to be involved in the regulation of trans-cellular water flow for long-distance transport in the root and leaf tissues. Their role is also critical for short-distance water transport and osmotic adjustments within a cell and between the cytoplasm and the cell wall space. AQP abundance is regulated developmentally in a cell-specific manner and by environmental signals. The activity of AQPs is also regulated by different post-translational regulation mechanisms, which provide an efficient way for rapid and reversible regulation of the water membrane permeability.
The research project aims at understanding the function and regulation of maize AQPs at the cellular level and in the whole plant subjected to various environmental conditions.

Aquaporin genes in maize
Screening of maize Expressed Sequenced Tags (ESTs) databases allowed us to identify 36 AQP cDNAs. A phylogenetic analysis of the maize AQPs showed that the distribution of AQP sequences between the four major subfamilies (PIP, TIP, NIP and SIP) was very similar in maize and Arabidopsis, suggesting that the separation into the different groups occurred before monocot-dicot divergence. However, a number of recent DNA duplication events arising after monocot-dicot separation could explain the presence of several closed isoforms within a species.

RNA and protein expression
To study the expression of maize AQP genes in different plant tissues, we developed quantitative and in situ RT-PCR approaches and prepared specific antibodies. We quantified and localized the expression of Zea mays plasma membrane PIP mRNA and proteins in primary maize root tip. Nearly all PIPs were expressed in primary roots. Expression was found to be dependent on the developmental stage of the root, with, in general, an increase in expression towards the elongation and mature zones.

Post-translational regulations
In addition to the modification of PIP expression, post-translational mechanisms appear to regulate PIP activity. We showed that non functional PIP1;2 and functional PIP2;1 physically interact in Xenopus oocytes leading to a higher amount of PIP1;2 in the plasma membrane (PM) and an increase in the osmotic water permeability. To study PIP localization and interactions in plant cells, we are currently using Fluorescence Resonance Energy Transfer technique.
We are also interested in determining the role of AQP phosphorylation on water permeability activity. Plasma membrane AQPs from etiolated maize shoots were in vivo and in vitro phosphorylated. This phosphorylation occurred on serine residues and was due to a Ca-dependent kinase associated with the plasma membrane.

Measurement of the osmotic water permeability of plant protoplasts
The understanding of the molecular and cellular mechanisms that underlie AQP regulation in plant cells requires accurate measurement of the osmotic water permeability by biophysical techniques. We developed a simple method to continuously measure and calculate the Pf of plant cells in which the protoplast cell volume and the osmolarity of the medium are both monitored in real time. The experimental data are analyzed by an off-line curve-fitting procedure, yielding relatively accurate Pf values over a large range of water permeability values

Key publications

Chaumont, F., Barrieu, F., Wojcik, E., Chrispeels, M.J. and Jung, R. (2001) Aquaporins constitute a large and highly divergent protein family in maize. Plant Physiol. 125, 1206-1215

Fetter, K., Van Wilder, V., Moshelion, M. and Chaumont, F. (2004) Interactions between plasma membrane aquaporins modulate their water channel activity. Plant Cell, 16, 215-228

Moshelion, M., Moran, N. and Chaumont, F. (2004) Dynamic changes in osmotic water permeability of cell membrane during an osmotic challenge. Plant Physiol. 135, 2301-2317

Chaumont, F., Moshelion, M. and Daniels, M.J. (2005) Regulation of plant aquaporin activity. Biology of the Cell, 97, 749-764

Hachez, C., Zelazny, E. and Chaumont, F. (2006) Modulating the expression of aquaporin genes in planta: a key to understand their physiological functions? Biochim. Biophys. Acta - Biomembranes, In press

Hachez, C., Moshelion, M., Zelazny, E., Cavez, D. and Chaumont, F. (2006) Localization and quantification of plasma membrane aquaporin expression in maize primary root: a clue to understanding their role as cellular plumbers. Plant Mol. Biol. In press