Guard-cell apoplastic sucrose concentration--a link between leaf photosynthesis and stomatal aperture size in the apoplastic phloem loader Vicia faba L.

In broad bean (Vicia faba L.), an apoplastic phloem loader, the sucrose concentration increases up to approximately 2 mM in the leaf apoplast and up to approximately 150 mM in the guard-cell apoplast during the photoperiod. This high concentration in the guard-cell apoplast results from transpiration and is sufficient osmotically to reduce stomatal aperture size by up to 3 microm or approximately 25% of the maximum aperture size. In this paper, we investigated a parallel and required role for high bulk-leaf apoplastic sucrose concentration, which correlates with high photosynthesis rate. An empirically determined combination of lowered light intensity and lowered CO(2) concentration reduced the photosynthesis rate to nominally one-fifth of the control value without a significant change in transpiration. This reduction in photosynthesis caused the sucrose concentration in the leaf apoplast--the immediate source pool for guard cells--to decrease by 70% (to 0.4 mM). In turn, sucrose concentration in the guard-cell apoplast decreased by approximately 80% (to approximately 40 mM). These results complete the required evidence for a non-exclusive, transpiration-linked, photosynthesis-dependent passive mechanism for the modulation of stomatal aperture size. In an ancillary investigation, hexoses in the bulk-leaf apoplast decreased when photosynthesis was lowered, but their concentrations in the guard-cell apoplast of control plants indicated that their osmotic contribution was negligible.

Guard cell apoplastic photosynthate accumulation corresponds to a phloem-loading mechanism.

Apoplastic phloem loaders have an apoplastic step in the movement of the translocated sugar, prototypically sucrose, from the mesophyll to the companion cell-sieve tube element complex. In these plants, leaf apoplastic sucrose becomes concentrated in the guard cell wall to nominally 150 mM by transpiration during the photoperiod. This concentration of external sucrose is sufficient to diminish stomatal aperture size in an isolated system and to regulate expression of certain genes. In contrast to apoplastic phloem loaders and at the other extreme, strict symplastic phloem loaders lack an apoplastic step in phloem loading and mostly transport raffinose family oligosaccharides (RFOs), which are at low concentrations in the leaf apoplast. Here, the effects of the phloem-loading mechanism and associated phenomena on the immediate environment of guard cells are reported. As a first step, carbohydrate analyses of phloem exudates confirmed basil (Ocimum basilicum L. cv. Minimum) as a symplastic phloem-loading species. Then, aspects of stomatal physiology of basil were characterized to establish this plant as a symplastic phloem-loading model species for guard cell research. [(14)C]Mannitol fed via the cut petiole accumulated around guard cells, indicating a continuous leaf apoplast. The (RFO+sucrose+hexoses) concentrations in the leaf apoplast were low, <0.3 mM. Neither RFOs (<10 mM), sucrose, nor hexoses (all, P >0.2) were detectable in the guard cell wall. Thus, differences in phloem-loading mechanisms predict differences in the in planta regulatory environment of guard cells.