The origin and composition of cucurbit "phloem" exudate

Abstract

Cucurbits exude profusely when stems or petioles are cut. We conducted studies on pumpkin (Cucurbita maxima) and cucumber (Cucumis sativus) to determine the origin and composition of the exudate. Morphometric analysis indicated that the exudate is too voluminous to derive exclusively from the phloem. Cold, which inhibits phloem transport, did not interfere with exudation. However, ice water applied to the roots, which reduces root pressure, rapidly diminished exudation rate. Sap was seen by microscopic examination to flow primarily from the fascicular phloem in cucumber, and several other cucurbit species, but primarily from the extrafascicular phloem in pumpkin. Following exposure of leaves to 14CO2, radiolabeled stachyose and other sugars were detected in the exudate in proportions expected of authentic phloem sap. Most of this radiolabel was released during the first 20 s. Sugars in exudate were dilute. The sugar composition of exudate from extrafascicular phloem near the edge of the stem differed from that of other sources in that it was high in hexose and low in stachyose. We conclude that sap is released from cucurbit phloem upon wounding but contributes negligibly to total exudate volume. The sap is diluted by water from cut cells, the apoplast, and the xylem. Small amounts of dilute, mobile sap from sieve elements can be obtained, although there is evidence that it is contaminated by the contents of other cell types. The function of P-proteins may be to prevent water loss from the xylem as well as nutrient loss from the phloem.

Figure 1.

Diagram of a transverse section of pumpkin stem adapted from Crafts (1932). The fascicular phloem (i.e. phloem in vascular bundles) is bicollateral, with both internal and external sieve tubes. The extrafascicular phloem inside the sclerenchyma ring is composed of peripheral sieve tubes closely adjacent to the vascular bundles, entocyclic sieve tubes just inside the sclerenchyma ring, and commissural sieve tubes that run transversely, linking these various elements to each other and to the fascicular phloem. Ectocyclic sieve tubes are found outside the sclerenchyma ring.

Figure 2.

Rate of exudation from the proximal cut surface of a pumpkin petiole (n = 3). The total volume exuded was 59.4 μL.

Figure 3.

Radiolabeled exudate from cut stems of pumpkin and cucumber. Leaf blades were exposed to ¹⁴CO₂ for 15 min, and 45 min later, stems were cut. Three representative experiments are shown. Charts on the left record exudation rate (A), ¹⁴C concentration (C), and ¹⁴C exudation rate (E) in cucumber and pumpkin. The inset in E records ¹⁴C exudation in a separate pumpkin experiment over a shorter time period. The three panels on the right record an experiment in which exudation rate (B) in a pumpkin plant declined, then rose again, then declined when ice water was applied to the soil (arrows).

Figure 4.

Volume (A) and ¹⁴C concentration (B) in exudate from the proximal and distal surfaces of a cut cucumber petiole. The leaf blade was exposed to ¹⁴CO₂ for 15 min, and the petiole was severed 45 min later.

Figure 5.

Suc and stachyose concentrations in exudate taken from the proximal and distal sides of cut cucumber petioles. Exudates were collected at 0.5 and 5 min after the petiole was severed. Samples from 10 to 15 petioles per plant were combined (n = 3 plants).

Figure 6.

Micrographs of exuding droplets. A, Cucumber vascular bundle stained with toluidine blue. The xylem (X) is stained blue, and other cells are stained purple. A small exuding droplet is seen over the external fascicular phloem (arrow). A larger droplet over the internal phloem (arrowhead) originated in the fascicular phloem but has grown to cover the extrafascicular phloem as well. B, Unstained pumpkin vascular bundle. An exuding droplet has emerged from the external fascicular phloem (arrow), which is uncommon in pumpkin. Another droplet is visible over the peripheral, internal extrafascicular phloem (arrowhead). The green region (double arrow) is the internal fascicular phloem. The white, crescent-shaped line is a diffraction image. C, Unstained pumpkin petiole section. Drops of exudate are visible over the ectocyclic phloem (arrow) and entocyclic phloem (arrowheads). Bars = 0.5 mm.

Figure 7.

Exudate sugar concentrations. Sugar concentrations are shown in cucumber (A and C) and pumpkin (B and D). In A and B, ice water was applied to the soil, and 10 min later, the stem was cut. After rapid blotting, exudate was collected from different phloem types. Data are from approximately 100 cuts per plant (n = 3 plants). In C and D, ice water was not applied to the soil. The stems were cut, and exudate from all phloem types was sampled together. Data are from three to five cuts per plant (n = 3 plants).

Figure 8.

¹⁴C in photosynthetically labeled sugars. Leaves of pumpkin (A) and cucumber (B) were exposed to ¹⁴CO₂ for 15 min. Forty-five minutes later, stems were cut and exudates were collected over two time periods. Three to five sampling cuts were made per plant. Radiolabeled sugars were analyzed by thin-layer chromatography (n = 3 plants).