Sorbitol is the predominant phloem-translocated carbohydrate in apple. The pathway—either apoplastic or symplastic—of sugar transport from photosynthetic cells to the phloem is not established. Furthermore, the presence of absence of phloem loading has not been tested. This study characterized the morphology and physiology of sugar movement to the phloem in apple leaves. An electron micrographic survey of apple leaf minor vein morphology was performed. Plasmodesmata were abundant and found at the interfaces of each cell type from mesophyll to sieve elements, indicating a symplastic sugar pathway. We also tested for a phloem loading mechanism. First, 14C-labeled sorbitol and sucrose were introduced exogenously to leaf discs to determine if they are loaded into veins from the apoplast. Although leaf discs floated on a solution containing either sugar actively accumulated label, the labeling pattern was diffuse, with no accumulation in minor veins. The addition of the sulfhydryl reagent PCMBS to the leaf disc assay inhibited sugar uptake. We also attempted plasmolysis of apple leaf sections to measure the solute concentration difference between photosynthetic mesophyll cells and cells of the minor vein phloem. Apple leaf pieces fixed in a solution containing 1.5 mol/kg osmoticum did not plasmolyze. We conclude that although active uptake of both sorbitol and sucrose takes place in apple leaves, apoplastic phloem-loading is absent. Considering the high sugar concentration and the symplastic connectivity among leaf cell types, we propose that sugars are instead enter the phloem after moving down—rather than against—a concentration gradient.
Raffinose family oligosaccharides (RFOs) perform several physiological functions in plants. In addition to accumulating during seed formation, raffinose and stachyose are translocated in the phloem and may accumulate in response to low temperatures, drought, or salt stress. Although the synthesis of galactinol, as mediated by galactinol synthase (GAS), is the first committed step in RFO formation, its expression patterns are poorly understood in most species. We have cloned and characterized the expression of two galactinol synthase gene family members in melon (Cucumis melo L. Cantalupensis Group). Both CmGAS1 and CmGAS2 are highly expressed in mature leaves. Galactinol synthase transcription in leaves was not upregulated by either water or low temperature stresses. Transcripts of CmGAS1 were present in developing melon seeds at a time coincident with the formation of raffinose and stachyose. Based on the GAS expression and RFO accumulation patterns, we propose that RFOs in melon function in carbon translocation and seed desiccation tolerance.
Sorbitol (d-glucitol) is the major end product of photosynthesis in apple (Malus domestica Borkh.), as well as the predominant phloem-translocated carbohydrate. The mechanism by which sorbitol is phloem-loaded for transport to heterotrophic sink tissues is unknown. We hypothesized that a plasma membrane-bound H+/sorbitol symporter mediates apoplastic phloem-loading of sorbitol. To discover genes potentially encoding sorbitol transporters, a cDNA library was constructed from mature `Gala' apple leaves. A homologous probe was synthesized via PCR with primers were designed against the cherry fruit sorbitol transporter, PcSot1, and using library lysate as template. From an initial plating of approximately 5 × 105 clones, twelve positives were identified after three rounds of hybridization screening. Following single-pass, 5' end sequencing, the clones were sorted into four contiguous sequences. One clone was chosen from each contig for complete sequencing. The four clones, provisionally named MdSOT1-4 (Malus domesitca Sorbitol Transporter), potentially encode full-length cDNAs for sorbitol transporters: Translated-BLAST searching (blastx) revealed that the open reading frames encode the complete Pfam sugar transporter domain, and the most significant alignments are with sequences encoding known- and putative polyol and sugar transporters.