LePLDβ1 activation and relocalization in suspension-cultured tomato cells treated with xylanase

Phospholipase D ( PLD) has been implicated in various cellular processes including membrane degradation, vesicular trafficking and signal transduction. Previously, we described a PLD gene family in tomato ( Lycopersicon esculentum) and showed that expression of one of these genes, LePLD beta 1, was induced by treatment with the fungal elicitor xylanase. To further investigate the function of this PLD, a gene-specific RNAi construct was used to knock down levels of LePLDb1 transcript in suspension-cultured tomato cells. Silenced cells exhibited a strong decrease in xylanase-induced PLD activity and responded to xylanase treatment with a disproportionate oxidative burst. Furthermore, LePLD beta 1-silenced cell-suspension cultures were found to have increased polyphenol oxidase activity, to secrete less of the beta-D-xylosidase LeXYL2 and to secrete and express more of the xyloglucan-specific endoglucanase inhibitor protein XEGIP. Using an LePLD beta 1-green fluorescent protein ( GFP) fusion protein for confocal laser scanning microscopy-mediated localization studies, untreated cells displayed a cytosolic localization, whereas treatment with xylanase induced relocalization to punctuate structures within the cytosol. Possible functions for PLD beta in plant-pathogen interactions are discussed.