Abstract
The biochemical pathway of stachyose synthesis was localized by immunocytochemical and 14C-labeling techniques in mature Cucurbita pepo L. leaves. Galactinol synthase (GaS; EC 2.4.1.123), the first unique enzyme in this pathway, was immunolocalized within the intermediary cells of minor veins in conventionally fixed and cryo-fixed, resin-embedded sections using polyclonal anti-GaS antibodies and protein A-gold. Intermediary cells are specialized companion cells with extensive symplastic connections to the bundle sheath. Gold particles were not seen over the non-specialized companion cells of larger veins or over intermediary cells in young leaves prior to the sink-source transition. In another approach to localization, radiolabel was measured in isolated mesophyll tissue and whole tissue of leaves that were lyophilized following a 90-s exposure to 14CO2. Mesophyll, obtained by abrasion of the leaf surface, contained labeled sucrose, galactinol, raffinose and stachyose. However, the latter three labeled compounds constituted a smaller proportion of the neutral fraction than in whole-tissue samples, which also contained minor veins. We conclude that synthesis of galactinol, raffinose, and stachyose occurs in both mesophyll and intermediary cells, predominantly the latter.
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Abbreviations
- GaS:
-
galactinol synthase
- SDS-PAGE:
-
sodium dodecyl sulfate-polyacrylamide gel electrophoresis
Referencess
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We thank John Pierce, Phillip Kerr, and Brace Schweiger for the gift of anti-GaS antibody and M.K. Kandasamy for helpful discussions. This research was supported by National Science Foundation grant DCB-9104159, U.S. Department of Agriculture Competetive Grant 90000854, and Hatch funds.
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Beebe, D.U., Turgeon, R. Localization of galactinol, raffinose, and stachyose synthesis in Cucurbita pepo leaves. Planta 188, 354–361 (1992). https://doi.org/10.1007/BF00192802
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DOI: https://doi.org/10.1007/BF00192802