The phloem sap of cucurbits is being used to identify NCAPs that function over long distances. Although the genes for these NCAPs can be identified using a proteomics approach, functional tests are difficult to perform as the cucurbits are rather recalcitrant to transformation. In addition, agroinfiltration experiments are very erratic thus limiting the use of this system for expression analyses. To circumvent this problem, we have developed a high throughput screen based on an infectious clone of the cucurbit-infecting Zuchini yellows mosaic potyvirus (ZYMV). As potyviruses encode for a polyprotein that is post-translationally processed into mature proteins, it can be used as a vector to over-express proteins of interest (Hsu et al., 2004). An engineered ZYMV strain that causes only mild symptoms on cucurbits (Lin et al., 2002) is being used to screen for dominant negative mutants. This is being conducted through an analysis of NCAP over-expression on plant growth and development. Initial experiments are based on NCAPP1 (Lee et al., 2003) and RNA-binding proteins that are carried in the phloem translocation stream. The next phase of this study will be to investigate the molecular determinants involved in mediating NCAP entry into and egress from the phloem translocation stream.

Infectious ZYMV clones were kindly provided by Prof. Shyi-Dong Yeh, Department of Plant Pathology, National Chung Hsing University, Taichung, Taiwan.