Aquaporins facilitate the uptake of soil water and mediate the regulation of root hydraulic conductivity (Lp_r) in response to a large variety of environmental stresses. Here, we use Arabidopsis (Arabidopsis thaliana) plants to dissect the effects of salt on both Lp_r and aquaporin expression and investigate possible molecular and cellular mechanisms of aquaporin regulation in plant roots under stress. Treatment of plants by 100 mm NaCl was perceived as an osmotic stimulus and induced a rapid (half-time, 45 min) and significant (70%) decrease in Lp_r, which was maintained for at least 24 h. Macroarray experiments with gene-specific tags were performed to investigate the expression of all 35 genes of the Arabidopsis aquaporin family. Transcripts from 20 individual aquaporin genes, most of which encoded members of the plasma membrane intrinsic protein (PIP) and tonoplast intrinsic protein (TIP) subfamilies, were detected in nontreated roots. All PIP and TIP aquaporin transcripts with a strong expression signal showed a 60% to 75% decrease in their abundance between 2 and 4 h following exposure to salt. The use of antipeptide antibodies that cross-reacted with isoforms of specific aquaporin subclasses revealed that the abundance of PIP1s decreased by 40% as early as 30 min after salt exposure, whereas PIP2 and TIP1 homologs showed a 20% to 40% decrease in abundance after 6 h of treatment. Expression in transgenic plants of aquaporins fused to the green fluorescent protein revealed that the subcellular localization of TIP2;1 and PIP1 and PIP2 homologs was unchanged after 45 min of exposure to salt, whereas a TIP1;1-green fluorescent protein fusion was relocalized into intracellular spherical structures tentatively identified as intravacuolar invaginations. The appearance of intracellular structures containing PIP1 and PIP2 homologs was occasionally observed after 2 h of salt treatment. In conclusion, this work shows that exposure of roots to salt induces changes in aquaporin expression at multiple levels. These changes include a coordinated transcriptional down-regulation and subcellular relocalization of both PIPs and TIPs. These mechanisms may act in concert to regulate root water transport, mostly in the long term (≥6 h).

Summary: Plant defence against pathogens is controlled by disease resistance (R) gene products that directly or indirectly detect specific pathogen effectors. Plant–pathogen interactions have been proposed to follow a co‐evolutionary arms‐race model where R genes are recent and evolve rapidly in response to structural changes in matching pathogen effectors. However, the longevity and extensive polymorphism of R genes studied were more consistent with balancing selection maintaining ancient and diverse R genes or alleles. In bread wheat (Triticum aestivum), the Pm3 locus confers race‐specific resistance to wheat powdery mildew (Blumeria graminis f.sp. triticii). Here we describe recently generated Pm3 resistance alleles that all derive from one susceptible allele, Pm3CS, which is widespread among hexaploid bread‐wheat lines. One group of four Pm3 resistance alleles shows few, clearly delimited, polymorphic sequence blocks of ancient origin, embedded in sequences identical to Pm3CS and possibly derived from gene conversion. A second group of three alleles differs from Pm3CS by only two to five mutations, all non‐synonymous, and all in the leucine‐rich repeat‐encoding region. Transient transformation experiments confirmed that Pm3 resistance specificities are based on one or few amino acid changes. The Pm3CS allele was found in wild tetraploid wheat, the ancestor of hexaploid bread wheat, specifically from southern Turkey, a region proposed to be the site of wheat domestication. Based on these data, we propose that the Pm3 resistance alleles were generated in agricultural ecosystems after domestication of wheat 10 000 years ago. The evolution of Pm3 alleles in wheat is best described by the model of evolved recycling, where novel genetic variation is integrated in plant populations together with recycling of old variation.

Proteins with nucleotide-binding site (NBS) and leucine-rich repeat (LRR) domains are major components of the plant immune system. They usually mediate resistance against a subgroup of races of a specific pathogen. For the allelic series of the wheat powdery mildew resistance gene Pm3, alleles with a broad and a narrow resistance spectrum have been described. Here, we show that a broad Pm3 spectrum range correlates with a fast and intense hypersensitive response (HR) in a Nicotiana transient-expression system and this activity can be attributed to two particular amino acids in the ARC2 subdomain of the NBS. The combined substitution of these amino acids in narrow-spectrum PM3 proteins enhances their capacity to induce an HR in Nicotiana benthamiana, and we demonstrate that these substitutions also enlarge the resistance spectrum of the Pm3f allele in wheat. Finally, using Bph14, we show that the region carrying the relevant amino acids also plays a role in the HR regulation of another coiled-coil NBS-LRR resistance protein. These results highlight the importance of an optimized NBS-‘molecular switch’ for the conversion of initial pathogen perception by the LRR into resistance-protein activation, and we describe a possible approach to extend the effectiveness of resistance genes via minimal targeted modifications in the NBS domain.