CmPP16 from Cucurbita maxima was cloned and the protein was shown to possess properties similar to those of viral movement proteins. CmPP16 messenger RNA (mRNA) is present in phloem tissue, whereas protein appears confined to sieve elements (SE). Microinjection and grafting studies revealed that CmPP16 moves from cell to cell, mediates the transport of sense and antisense RNA, and moves together with its mRNA into the SE of scion tissue. CmPP16 possesses the characteristics that are likely required to mediate RNA delivery into the long-distance translocation stream. Thus, RNA may move within the phloem as a component of a plant information superhighway.

SummaryPlasmodesmata and the nuclear pore complex (NPC) mediate the selective trafficking of proteins and protein-nucleic acid complexes. The events underlying the translocation of endogenous and viral proteins through plasmodesmata were investigated to further explore the parallels between these cell-to-cell and intracellular communication systems. Studies performed with crosslinked KNOTTED1 (KN1) revealed that a conformational change is required for the cell-to-cell movement of this protein. Microinjection of gold-conjugated KN1 established that, as with the NPC, a combination of protein unfolding and microchannel dilation appears to be involved in protein translocation. However, during this process the extent of microchannel dilation is much less than observed for the NPC, which may reflect a physical limitation imposed by the cell wall. Co-injection of KN1-gold with unbound KN1 or cucumber mosaic virus movement protein (CMV-MP) established that the KN1-gold probe is highly effective at blocking plasmodesmal transport of KN1 and CMV-MP. This result provided the foundation for competition experiments which demonstrated that KN1 and the viral movement proteins of CMV and tobacco mosaic virus likely utilize a common receptor in the pathway for cell-to-cell transport of proteins. A combination of biochemical fractionation methods, an in vitro binding assay founded on the high affinity between KN1-gold and the putative common plasmodesmal receptor, and microinjection techniques were used to isolate plasmodesmal constituents involved in cell-to-cell transport. A model describing the steps involved in protein transport through plasmodesmata is presented.

Although no signs of toxicity were found after the prolonged presence of TB at a concentration of 0.06% in the vitreous cavity of rabbit eyes, marked damage occurred in the lower retina of 0.2% TB-treated eyes. The short-term presence of TB at a concentration of 0.06% in the vitreous cavity is harmless to the rabbit eye but a higher concentration of TB could be unsafe.

We report on the molecular, biochemical, and functional characterization of Cucurbita maxima phloem serpin-1 (CmPS-1), a novel 42-kDa serine proteinase inhibitor that is developmentally regulated and has antielastase properties. CmPS-1 was purified to near homogeneity from C. maxima (pumpkin) phloem exudate and, based on microsequence analysis, the cDNA encoding CmPS-1 was cloned. The association rate constant (k a ) of phloem-purified and recombinant His 6 -tagged CmPS-1 for elastase was 3.5 ؎ 1.6 ؋ 10 5 and 2.7 ؎ 0. with a 3:2 molar ratio. In vivo feeding assays conducted with the piercing-sucking aphid, Myzus persicae, established a close correlation between the developmentally regulated increase in CmPS-1 within the phloem sap and the reduced ability of these insects to survive and reproduce on C. maxima. However, in vitro feeding experiments, using purified phloem CmPS-1, failed to demonstrate a direct effect on aphid survival. Likely roles of this novel phloem serpin in defense against insects/ pathogens are discussed.The phloem long-distance translocation system of plants appears to function both as a nutrient delivery system and as an information superhighway (1-3). A central role for the phloem in the translocation of nutrients has long been recognized. The presence of plant hormones in the phloem sap (4, 5) implicated this long-distance transport pathway in the delivery of signaling molecules. Recent studies provided new insights into the nature of the information molecules being transported from mature leaves, via the phloem, to distant plant organs. Irrefutable evidence has been obtained for the translocation of certain proteins (6 -9). In addition, it has also been demonstrated that specific RNA molecules are present in the phloem sap (10, 11) and some move to distant tissues, where they appear to influence post-transcriptional events (9, 12).Given the importance of this nutrient/information delivery system to the functioning of the plant, it was axiomatic that plants had to evolve mechanisms to protect the operational integrity of the phloem. Maintenance of structural integrity required the development of systems able to rapidly respond to physical damage, imposed either by environmental forces or herbivory; the sealing of disrupted sieve tubes involves deposition of material at the level of the sieve plate pore (1). In the enucleate sieve tube system of angiosperms, maintenance of membrane integrity has also been transferred to the neighboring companion cells and likely involves the delivery of essential constituents via plasmodesmata (6,13,14).To ensure the integrity of the signaling components, the plant needs also to protect against protein and RNA degradation occurring within the phloem sap. This capacity appears to have been achieved through the development of a control system that regulates the plasmodesmal-mediated exchange of macromolecules between companion cells and the sieve tube system (9, 12, 13). The absence of proteinase activity (15) within the phloem sap is consistent with this mo...