ISOLATION, CHARACTERIZATION AND GENETIC MANIPULATION OF XYLELLA FASTIDIOSA HEMAGGLUTININ GENES

• Author(s): Kirkpatrick, Bruce; Bruening, George; Feldstein, Paul; Igo, Michele; Voegel, Tanja; Warren, Jeremy;
• Abstract: Xylella fastidiosa (Xf) possesses genes for hemagglutinins (HAs), large adhesion proteins involved in cell-cell aggregation and biofilm formation. Mutations in either one of the functional HAs, HxfA (PD2118) or HxfB (PD1792), result in hypervirulent strains that move faster and cause more severe disease in grapevines. Computer analyses of the HA proteins identified several regions that might be possible adhesion domains (ADs) responsible for cell-cell and/or cell-surface binding. We cloned 6 Xf HA fragments that may contain potential ADs into protein expression vectors and to date have prepared antibodies against 1 AD protein fragment that is conserved in both HxfA and B. Recombinant proteins from the other 5 ADs are being purified and prepared for injection. Western blot analyses of using Xf proteins extracted from Xf cells grown in liquid culture showed a very faint reaction with an Xf protein of approximately 220kd but this result needs to be confirmed using a higher quality antibody. Recent discoveries in the Bruening lab using phage technology indicate that HA are more abundant in cells grown on solid medium compared to liquid medium and we are now repeating our Western analyses using cells grown on solid medium. Once Xf HA cell-cell binding domains are identified they will be expressed in transgenic tobacco and grapevines where we hope the proteins will act as a molecular glue to aggregate insect-inoculated Xf cells, retard their ability to systemically colonize plants and potentially provide a unique form of resistance against PD.
• Publication Date: Nov 2006
• Journal: 2006 Pierce's Disease Research Symposium