POTATO GUARD-CELLS RESPOND TO DRYING SOIL BY A COMPLEX CHANGE IN THE EXPRESSION OF GENES RELATED TO CARBON METABOLISM AND TURGOR REGULATION

Altering stomatal function by a guard cell-targeted transgenic approac h with the aim of increased stress tolerance and crop yield requires k nowledge of the natural fluctuations of stomatal gene expression under stress conditions. We developed a fast method for the isolation of RN A from epidermal fragments of potato leaves (Solanum tuberosum L. cv. Desiree), demonstrated that this RNA preparation is highly enriched in guard cell transcripts and used this method to investigate the respon se of gene expression in guard cells to mild drought stress. Drought w as applied in planta by withholding water over a period of 2-4 days. i n the following work responses observed under these conditions are cal led 'long-term' in contrast to immediate (short term) stomatal opening and closing responses to environmental stress. We observed both gene- specific increases and decreases of steady-state transcript levels. In particular, the mRNA levels of sucrose synthase and sucrose-phosphate synthase were elevated 5.5-fold and 1.4-fold, respectively. In contra st, expression of an inwardly rectifying K+ channel from guard cells ( kst1) and of a plasma membrane H+-ATPase (pha2) was reduced to 26% and 36%, respectively, of the expression in watered controls. In addition , expression of vacuolar invertase, UDP-glucose pyrophosphorylase, ADP -glucose pyrophosphorylase (large subunit), cytosolic glyceraldehyde-3 -phosphate dehydrogenase, a sucrose/H+ cotransporter, and a novel isof orm of phosphoenolpyruvate carboxylase were also reduced. Other genes exhibited unaltered expression. Compared with the response in whole le aves, the transcript levels of phosphoenolpyruvate carboxylase, vacuol ar invertase, and cytosolic glyceraldehyde-3-phosphate dehydrogenase w ere regulated guard cell specifically. Most importantly, changes in st eady-state transcript levels were complete before the onset of a decre ase in leaf water potential, when drought-induced stomatal closure was already obvious. These data support the hypothesis that a systemic dr aught-stress signal acts not only on short-term stomatal movements but also on long-term gene expression in guard cells. Such long-term chan ges in gene expression might contribute to the fine-tuning of guard ce ll responses to environmental stimuli.