1.11.29
HOST SPECIES-DEPENDENT AND VIRUS STRAIN-DEPENDENT LONG-DISTANCE MOVEMENT OF POTATO A POTYVIRUS

J ANDREJEVA1,2, A MERITS1, Ü PUURAND1, L JÄRVEKÜLG2, M SAARMA1 and JPT VALKONEN1,3

1Institute of Biotechnology, Viikki Biocenter 1, PO Box 56, FIN-00014, University of Helsinki, Finland; 2Institute of Chemical Physics and Biophysics, Akadeemia tee 23, Tallinn, Estonia EE0026; 3Department of Plant Biology, Genetic Centre, Swedish University of Agricultural Sciences (SLU), PO Box 7080, S-75007 Uppsala, Sweden

Background and objectives
Systemic or long-distance spread of plant viruses involves at least three steps: virus replication in initially infected cells; cell-to-cell movement via plasmodesmata; and long-distance transport, after which virus infects cells and organs distant from the site of initial infection. Recent evidence suggests that two viral-encoded proteins - coat protein (CP) and helper component proteinase (HC-Pro) - are responsible for systemic movement of potyviruses [1]. The object of our research is potato A potyvirus. It infects potato plants all over the world, reducing yields up to 40%. Lack of systemic spread of one PVA isolate (PVA-B11) was observed in several potato cultivars [2], however, PVA-B11 replicates in inoculated potato leaves. PVA isolate U (PVA-U) is able to move systemically in potato plants and, compared to PVA-B11, it has four amino-acid substitutions in HC-Pro and seven amino-acid substitutions in the N-terminal part of the CP. The aim of our study was to restore systemic movement of PVA-B11 in potato plants using the full-length, infectious cDNA clone of PVA-B11 [3] as a target for site-directed mutagenesis and gene exchange.

Results and conclusions
Three types of PVA-B11 mutants were constructed: (i) CP gene of PVA-B11 was replaced with CP gene of PVA-U; (ii) several nucleic acid substitutions were introduced to PVA-B11 HC-Pro to make it resemble the HC-Pro of PVA-U; and (iii) both HC-Pro and CP of PVA-B11 were replaced with those of PVA-U. All three constructs were infectious and spread systemically on tobacco plants (Nicotiana tabacum cv. Samsun). The first two mutants mentioned above and PVA-B11 also replicated in the inoculated leaves of potato cv. Pentland Ivory, and were detected by ELISA. However, the third mutant with the double gene replacement did not give a positive response by ELISA. No infection was detected with any three mutants or PVA-B11 in the non-inoculated, upper leaves, in contrast to PVA-U. Our data suggest that HC-Pro and CP of PVA are not sufficient alone to provide PVA-B11 with the capacity to move over long distance in potato cv. Pentland Ivory.

References
1. Cronin S, Verchot J, Haldeman-Cahill R et al., 1995. Plant Cell 7, 549-559.
2. Valkonen JPT, Puurand Ü, Slack SA et al., 1995. Plant Disease 79, 748-753.
3. Puurand Ü, Valkonen JPT, Mäkinen K et al., 1996. Virus Research 40, 135-140.