1. ABA increases [Ca 2+ ] cyto influx through plasma membrane or release from internal compartments Verapamil: an inhibitor of extracellular Ca 2+ channels TMB-8: an an inhibitor of intracellular Ca 2+ channels J. Exp. Bot. (1995) 46: 973-979

2. Microinjected Ca 2+ (cyto) sensitive dye- fura-2

3. Non-invasive method: transgenetic plants expressing the gene for the calcium indicator protein yellow cameleon ABA-induced calcium oscillation in guard cells of Arabidopsis

4. Web Topic 23.9 A Ca 2+ binding fusion protein: Ca 2+ indicator yellow cameleon Composed 4 domains: enhanced cyan fluorescence protein (ECFP) enhanced yellow fluorescence protein (EYFP) calmodulin (CaM) CaM-binding protein (M13) The 535/480 emission ratio  [Ca 2+ ] cyto

5. * p. 606 Reactive oxygen species (ROS), inositol 1,4,5-triphosphate (IP 3 ), cyclic ADP-ribose (cADPR), self-amplifying  activate channel * ABA  nitric oxide  cADPR  stomatal closure * p. 607R Auxin-induced stomatal opening  [Ca 2+ ] cyto   the location and periodicity of calcium oscillations rather than the overall concentration of cytosolic calcium, determine the cellular response. Phalaenopsis: citric acid/ malate

6. ¤ ABA inhibits the plasma membrane H + -ATPase also contribute membrane depolarization  indirectly ABA inhibition proton pump is via increase [ca 2+ ] cyto and alkalization of the cytosol (pH 7.67  7.94, p. 607R) p. 452

7. ¤ p. 603 ABA activate slow and rapid (06) anion channels in guard cells  large amounts of Cl - and malate 2- to escape from the cell  depolarization (more positive) / calcium factor  open voltage-gated K + efflux channels ( long-term membrane depolarization) Anion channel inhibitors: 5-nitro-2,3-phenylpropylaminobenzoid (NPPB) – slow type block ABA-induced stomatal closing 4,4’-diisothiocyanatostilbene-2,2’-disulfonic acid (DIDS) – rapid type have no (02) effect on ABA-induced stomatal closing

8. ¤ ABA prevents light-induced stomatal opening – inhibit the inward K + channel ¤ stomatal closing of guard cells under stress prevent opening: inhibit inward K + channels and plasma membrane H + pumps promote closing: activate outward anion channels, then active K + efflux channels  ABA signaling involves both Ca 2+ dep- and independent pathway  “network robustness”

9. Daily changes in stomatal aperture, and in sucrose and K + content Red light: only sucrose accumulation Stomatal opening is associated primarily with K + uptake, and closing is associated with a decrease in sucrose content, why? Plant cell (04) 16: 1143-1162 The ascorbic acid redox state controls guard cell signaling and stomatal movement