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Raffinose oligosaccharide concentrations measured in individual cell and tissue types in Cucumis melo L. leaves: implications for phloem loading

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Abstract

Raffinose, stachyose, and galactinol are synthesized in intermediary cells (specialized companion cells) of the minor-vein phloem of cucurbits. To better understand the role of these carbohydrates and the regulation of their synthesis and transport, we measured the concentrations of each of the components of the raffinose oligosaccharide synthetic pathway in mesophyll and sieve element-intermediary cell complexes (SE-ICCs) in the leaves of melon (Cucumis melo L. cv. Hale's Best Jumbo). These concentrations are consistent with a polymer-trapping mechanism for phloem loading, with sucrose diffusing from mesophyll into intermediary cells and being made into raffinose and stachyose, which are too large to diffuse back to the mesophyll. To determine carbohydrate concentrations, we developed a method involving microdissected tissues. Blind endings of areoles, and mesophyll surrounding these veins, were separately removed from lyophilized leaf tissue. Carbohydrates were quantitated by high-performance liquid chromatography with pulsed amperometric detection. A small amount of mesophyll remained attached to the blind endings; the carbohydrate contribution of these cells to the vein sample was eliminated by subtraction, based on the amount of chlorophyll. Volumes of cells and subcellular compartments were calculated by morphometric analysis and were used to calculate carbohydrate concentrations. Assuming no subcellular compartmentation, the additive concentration of sugars in the SE-ICCs of minor veins is about 600 mM. Stachyose and raffinose concentrations are about 330 mM and 70 mM, respectively, in SE-ICCs; concentrations of these sugars are much lower in mesophyll (0.2 and 0.1 mM). This is consistent with the view that stachyose and raffinose are unable to pass through the plasmodesmata between intermediary cells and bundle-sheath cells. Sucrose levels appear to be higher in the SE-ICC (about 130mM) than in the mesophyll (about 10 mM), but if compartmentation is taken into account the gradient for sucrose is probably downhill from mesophyll to intermediary cells. Flux through plasmodesmata between the bundle sheath and intermediary cells was calculated and was found to be within the range of values of flux through plasmodesmata reported in the literature.

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Abbreviations

BS-IC:

bundle sheath-intermediary cell

PC:

plasmodesmatal channel

SE-ICC:

sieve element-intermediary cell complex

SEL:

size exclusion limit

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Authors and Affiliations

  1. Section of Plant Biology, Cornell University, 14853, Ithaca, NY, USA

    Edith Haritatos & Robert Turgeon

  2. Institute of Plant Biology, University of Zürich, Zollikerstrasse 107, CH-8008, Zürich, Switzerland

    Felix Keller

Authors
  1. Edith Haritatos
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  2. Felix Keller
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  3. Robert Turgeon
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Correspondence to Edith Haritatos.

Additional information

We would like to thank Gayle Volk, Philip Laible, Canan Inan, Esther Gowan, Richard Medville, Nathan Wilson, Jessica Plant, and Steven Boese for their help, Thomas Owens, M.V. Parthasarathy, and Ian Merwin for use of equipment, and Nancy Haritatos for suggestions. This research was supported by U.S. Department of Agriculture Competitive Grant 94-37306-0351 (R.T.), the Swiss National Foundation (F.K.), and a NSF/DOE/USDA Cornell Plant Science Center fellowship (E.H.).

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Haritatos, E., Keller, F. & Turgeon, R. Raffinose oligosaccharide concentrations measured in individual cell and tissue types in Cucumis melo L. leaves: implications for phloem loading. Planta 198, 614–622 (1996). https://doi.org/10.1007/BF00262649

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