Proceedings of the XLV Italian Society of Agricultural Genetics - SIGA Annual Congress
Salsomaggiore Terme, Italy - 26/29 September, 2001
ISOLATION, GENOME ORGANIZATION AND EXPRESSION OF AQUAPORIN-ENCODING GENES FROM THE MARINE PHANEROGAM POSIDONIA OCEANICA
MAESTRINI P., NATALI L., GIORDANI T., CAVALLINI A.
Dipartimento di Biologia delle Piante Agrarie, Sezione di Genetica, Via Matteotti 1/B, 56124 Pisa
aquaporin, Posidonia oceanica, water transport
The diffusion of water through terrestrial plants can be driven by concentration gradients of osmotically active solutes or by physical pressure, generating an osmotic or hydrostatic force, respectively. Beyond simple diffusion across a lipid bilayer, the existence of proteinaceous water channels, aquaporins, in plant membranes has been established. Aquaporins belong to a highly conserved group of membrane major intrinsic proteins (MIPs) with molecular masses of 26 to 30 kDa. Within a species, aquaporin-encoding genes form a large family, for example, more than 30 MIPs have been found in Arabidopsis, with specificity in function and organ, tissue, and cellular localization (Johansson et al., 2000).
Aquaporins are likely to be important for both the whole plant (for water transport to and from vascular tissues) and the cells (for buffering osmotic fluctuations in the cytosol). Putative aquaporin-encoding DNA sequences have been isolated from a number of terrestrial plants, where passive water flux through membrane channels is determined by leaf transpiration and water absorption by roots.
No aquaporins have been until now isolated from marine plants, where physical forces that regulate water flux should be, expecially at leaf level, very different from those operating in terrestrial plants, since water loss from leaves is probably not caused by transpiration but by osmotic forces.
Among marine plants, Posidonia oceanica is a monocotyledon growing along coastal waters of the Mediterranean basin and form dense infralittoral populations that frame the so-called Posidonia meadow ecosystem (Boudouresque and Meinesz, 1982). Because of its high productivity, P. oceanica represents an important food substrate for many marine organisms (Novak, 1982).
In our experiments, we isolated, by RT-PCR and RACE (Rapid Amplification of cDNA Ends), two complete cDNAs putatively encoding aquaporins. The first coding sequence, PIP1a, is 867 bp long and shows high similarity to plasma membrane aquaporin of other plants species; the second, TIP1, 753 bp long, is highly similar to tonoplast aquaporins. The two sequences were studied after isolation of the corresponding genes by PCR on genomic DNA: PIP1a is interrupted by two introns, TIP1 by one intron. Southern analysis showed that these sequence belong to a gene family. Finally, the expression of these genes was studied, by northern blot and hybridization, in different tissues of P. oceanica, in response to changes in seawater salinity and in response to mercury exposure, a metal typically blocking aquaporin-based pores.