Eriko M. Miura 1 , Byung-Chun Yoo 2 , and William J. Lucas 1

There is now accumulating evidence that supports the concept of RNA as long-distance information macromolecule (Lucas et al., 2001).   One of the projects in our laboratory focuses on the identification of molecular components that underlie the selective trafficking of RNA via the phloem.   Studies on plant viral movement proteins and intracellular RNA localization in animal systems have shown that RNA moves as a ribonucleoprotein (RNP) complex (Gilbertson et al ., 1996; Grünert et al. , 1996).   RNA-binding proteins (RBPs) are important components of such complexes, as they combine the ability to recognize specific RNA with the capacity to interact with other proteins.   In our current working model describing long-distance RNA trafficking, a non-cell autonomous RNA originating in the nucleus of a companion cell is bound by specific RBPs and their interaction partners to form a RNP complex, which then is targeted to the plasmodesmata (PD) at companion cell-sieve element interface to enter the phloem translocation stream.   Upon reaching the target tissue (e.g., shoot apex), the RNP complex is then unloaded from the sieve-tube system, again via specific interactions with the PD.   To elucidate the components of potentially complex machinery involved in RNA trafficking, we devised a Northwestern screen using fractionated pumpkin phloem sap and a radiolabeled phloem-derived RNA probe.    We were able to detect ~40 potential RBPs in this manner.   Of these proteins, a 17kD protein (CmPP17, or Cucurbita maxima phloem protein 17kD) showed high affinity for dsRNA substrate, and was later identified to be a homolog of eukaryotic initation factor 5A (eIF-5A).   Current literature on eIF-5A supports the role of the protein as a nucleocytoplasmic RNA shuttle (Lipowsky et al., 2000), but its physiological function remains elusive . Here, we report our progress on characterization of CmPP17.   Work is currently ongoing to identify the CmPP17 ortholog among the 3 members of eIF-5A gene family in Arabidopsis .     

Possible functions of plant eIF-5A:

1. Currently favored hypothesis of eIF-5A function is that it is a nucleocytoplasmic RNA shuttle. Given the conservation of eIF-5A in all organisms except for prokaryotes, it is possible that plants express at least one isoform of eIF-5A with a conserved function that acts cell-autonomously. Such an isoform would be expected to recognize a subset of RNA which then is targeted to the polysomes.
2. Here, a CmPP17-like eIF-5A is depicted as an RNA shuttle protein whose function has been extended to the CC-SE (companion cell-sieve element) interface as part of the long-distance macromolecular trafficking pathway. A subset of RNA with specific zip-code (PSE) is recognized by eIF-5A, which is then trafficked to the CC-SE interface to access the phloem translocation stream. eIF-5A then can be shuttled back to the CC, or it can traffic long-distance with the RNP complex to the distal target tissue.
3. Another possibility is that the CmPP17-like eIF-5A isoform functions exclusively at the CC-SE interface.