Abscisic acid regulates root hydraulic conductance via aquaporin expression modulation in Nicotiana tabacum.

Abscisic acid (ABA) modifies the hydraulic properties of roots by increasing root water flux. The effects of ABA on aquaporin content and root hydraulic conductance are controversial. We addressed these effects via a combination of experiments. Tobacco (Nicotiana tabacum) plants were grown hydroponically, and ABA (1 microM) was exogenously applied to the roots. Then, the water transport properties of tobacco roots and expression of PIP-type aquaporins were examined. ABA increased the sap flow rate (Jv) and also the osmotic root hydraulic conductance (Lp(r-o)) of excised tobacco roots after 24h. The expression of three aquaporin PIP-type genes and PIP1s proteins abundance in tobacco roots were analyzed by real-time PCR and protein gel blot analysis, respectively. Interestingly, the accumulation of NtAQP1, NtPIP1;1 and NtPIP2;1 transcripts and NtPIP1;1 and NtAQP1 proteins abundance was significantly increased. Although the antibody used recognize NtPIP1;1 and NtAQP1, most probably it also recognizes other PIP1 proteins present in tobacco. Thus, the increase in the expression of the three PIP-type genes and other PIP1s proteins abundance caused by ABA were correlated with an increase in Lp(r-o) and Jv. ABA therefore facilitated the cell-to-cell component of water transport across the root cylinder. The subcellular localization of NtPIP1;1- and NtPIP2;1-GFP was investigated by protoplast transformation with chimeric gene, showing NtPIP2;1 localization in plasma membrane and NtPIP1;1 retention in the endoplasmic reticulum (ER). However, ABA did not change subcellular localization of NtPIP1;1 from ER to plasma membrane.

Drought stress alters water relations and expression of PIP-type aquaporin genes in Nicotiana tabacum plants.

Plasma membrane intrinsic proteins (PIPs), a type of aquaporins, mediate water transport in many plant species. In this study, we investigated the relationship between the functions of PIP-type water channels and water relations of tobacco plants (Nicotiana tabacum cv. Samsun) under drought stress. Drought stress treatments have led to reductions in the stomatal conductance, transpiration, water potential and turgor pressure in leaves, and also the sap flow rate and osmotic hydraulic conductance in roots. In contrast, leaf osmotic pressure was increased in response to drought stress. Interestingly, the accumulation of NtPIP1;1 and NtPIP2;1 transcripts was significantly decreased, but only that of the NtAQP1 transcript was increased under drought stress. Functional analysis using Xenopus laevis oocytes revealed that NtPIP2;1 shows marked water transport activity, but the activities of NtAQP1 and NtPIP1;1 are weak or almost negligible, respectively, when expressed alone. However, co-expression of NtPIP1;1 with NtPIP2;1 significantly enhanced water transport activity compared with that of NtPIP1;1- or NtPIP2;1-expressing oocytes, suggesting that these two aquaporins may function as a water channel, forming a heterotetramer. Heteromerization of NtPIP1;1 and NtPIP2;1 was also suggested by co-expression analyses of NtPIP1;1-GFP (green fluorescent protein) and NtPIP2;1 in Xenopus oocytes. Re-watering treatments recovered water relation parameters and the accumulation of the three NtPIP transcripts to levels similar to control conditions. These results suggest that NtPIP1;1 and NtPIP2;1 play an important role in water transport in roots, and that expression of NtPIP1;1 and NtPIP2;1 is down-regulated in order to reduce osmotic hydraulic conductance in the roots of tobacco plants under drought stress.