Abstract
Aquaporins are membrane channels that facilitate the transport of water and other small molecules across the cellular membranes. We examined the role of six aquaporins of Vitis vinifera (cv. Touriga nacional) in the transport of water and atypical substrates (other than water) in an aqy-null strain of Saccharomyces cerevisiae. Their functional characterization for water transport was performed by stopped-flow fluorescence spectroscopy. The evaluation of permeability coefficients (Pf) and activation energies (Ea) revealed that three aquaporins (VvTnPIP2;1, VvTnTIP1;1 and VvTnTIP2;2) are functional for water transport, while the other three (VvTnPIP1;4, VvTnPIP2;3 and VvTnTIP4;1) are non-functional. TIPs (VvTnTIP1;1 and VvTnTIP2;2) exhibited higher water permeability than VvTnPIP2;1. All functional aquaporins were found to be sensitive to HgCl2, since their water conductivity was reduced (24–38%) by the addition of 0.5 mM HgCl2. Expression of Vitis aquaporins caused different sensitive phenotypes to yeast strains when grown under hyperosmotic stress generated by KCl or sorbitol. Our results also indicate that Vitis aquaporins are putative transporters of other small molecules of physiological importance. Their sequence analyses revealed the presence of signature sequences for transport of ammonia, boron, CO2, H2O2 and urea. The phenotypic growth variations of yeast cells showed that heterologous expression of Vitis aquaporins increased susceptibility to externally applied boron and H2O2, suggesting the contribution of Vitis aquaporins in the transport of these species.
Highlights
Aquaporins play a crucial role in maintaining water and ion homeostasis of plants, essential for plant cell integrity, growth and survival in their ever-changing environment
Aquaporins belong to Major Intrinsic Proteins (MIPs) family and based on their sequence similarity and sub-cellular localization, plant aquaporins are divided in seven subfamilies: the plasma membrane intrinsic proteins (PIPs), the tonoplast intrinsic proteins (TIPs), the nodulin-26-like intrinsic proteins (NIPs), the small intrinsic proteins (SIPs), the GlpF-like intrinsic proteins (GIPs), the hybrid intrinsic proteins (HIPs) and the uncategorized X intrinsic proteins (XIPs) [2]
Touriga nacional in this study were clearly identified in the genome of Pinot noir variety and three of them were clustered in their subsequent groups of PIPs, while other three in TIPs subfamilies (Figure 1)
Summary
Aquaporins play a crucial role in maintaining water and ion homeostasis of plants, essential for plant cell integrity, growth and survival in their ever-changing environment. These water channels can provide rapid and reversible changes to cells hydraulic conductance by modulating membrane water permeability [1]. Studies on plant aquaporins revealed their role far beyond the membrane water transport. Besides water, they are reported to transport other small molecules and/or gases of physiological importance (reviewed by [3]), suggesting their versatile functions in plants. Putative substrate specificities of aquaporins are generally assigned by the presence of specific amino acid residues at well-defined positions in the sequences [4]
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