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. 1997 Dec 9;94(25):14150-5.
doi: 10.1073/pnas.94.25.14150.

Phloem sap proteins from Cucurbita maxima and Ricinus communis have the capacity to traffic cell to cell through plasmodesmata

S Balachandran  1 , Y Xiang, C Schobert, G A Thompson, W J Lucas
Affiliations

Phloem sap proteins from Cucurbita maxima and Ricinus communis have the capacity to traffic cell to cell through plasmodesmata

S Balachandran et al. Proc Natl Acad Sci U S A. .

Abstract

In angiosperms, the functional enucleate sieve tube system of the phloem appears to be maintained by the surrounding companion cells. In this study, we tested the hypothesis that polypeptides present within the phloem sap traffic cell to cell from the companion cells, where they are synthesized, into the sieve tube via plasmodesmata. Coinjection of fluorescently labeled dextrans along with size-fractionated Cucurbita maxima phloem proteins, ranging in size from 10 to 200 kDa, as well as injection of individual fluorescently labeled phloem proteins, provided unambiguous evidence that these proteins have the capacity to interact with mesophyll plasmodesmata in cucurbit cotyledons to induce an increase in size exclusion limit and traffic cell to cell. Plasmodesmal size exclusion limit increased to greater than 20 kDa, but less than 40 kDa, irrespective of the size of the injected protein, indicating that partial protein unfolding may be a requirement for transport. A threshold concentration in the 20-100 nM range was required for cell-to-cell transport indicating that phloem proteins have a high affinity for the mesophyll plasmodesmal binding site(s). Parallel experiments with glutaredoxin and cystatin, phloem sap proteins from Ricinus communis, established that these proteins can also traffic through cucurbit mesophyll plasmodesmata. These results are discussed in terms of the requirements for regulated protein trafficking between companion cells and the sieve tube system. As the threshold value for plasmodesmal transport of phloem sap proteins falls within the same range as many plant hormones, the possibility is discussed that some of these proteins may act as long-distance signaling molecules.

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Figures

Figure 1
Figure 1
Phloem exudate collected from the stem of 2-month-old C. maxima plants for use in microinjection studies. Shown is the protein pattern obtained by SDS/PAGE (lane 1). Lane 2 displays the molecular weight markers. Distribution of phloem proteins present in size-fractionated samples are evident: fraction I, 10–20 kDa; fraction II, 20–25 kDa; fraction III, 25–50 kDa; fraction IV, 50–80 kDa; fraction V, 80–100 kDa; fraction VI, 100–200 kDa. Individual proteins used in microinjection experiments are indicated by darts. UBI, ubiquitin.
Figure 2
Figure 2
Microinjection experiments performed to determine the threshold level of PP2 required to potentiate cell-to-cell movement of coinjected 20-kDa FITC-dextran. PP2 used in these experiments was either purified from cucurbit phloem exudate or expressed in and extracted from E. coli (Inset). In each microinjection series, the level of FITC-dextran was held constant (1 mM) whereas the level of PP2 was serially reduced until fluorescence associated with the FITC-dextran did not move out from the injected cell.
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