EXPRESSION OF PHOSPHOLIPASE-D DURING CASTOR BEAN LEAF SENESCENCE

Membrane deterioration in plant senescence is commonly associated with progressive decreases in membrane phospholipid content. This study in vestigated the expression and regulation of phospholipase D (PLD; EC 3 .1.4.4) during senescence in castor bean (Ricinus communis L. cv Hale) leaf discs. The rate of leaf senescence was accelerated by 50 mu M ab scisic acid and was attenuated by 50 mu M cytokinin during incubation at 23 degrees C for up to 5 d. Leaf senescence was indicated by decrea ses in the content of total proteins, chlorophyll, and phospholipids. PLD activity in both membrane-associated and cytosolic fractions showe d a gradual increase in the absence of phytohormones. Abscisic acid st imulated an increase in membrane-associated PLD and had little effect on the soluble form. On the other hand, cytokinin retarded the increas e in membrane-associated PLD. Immunoblotting analysis using PLD-specif ic antibodies revealed that the changes in PLD activity were correlate d with those of PLD protein. Analysis of PLD by nondenaturing PACE sho wed the appearance of a PLD structural variant, PLD 3, in abscisic aci d-treated leaf discs. Northern blotting analysis using a PLD cDNA prob e revealed an increase in PLD mRNA in senescing leaf discs. These data indicate complex mechanisms for the regulation of PLD during senescen ce, which include increases in membrane-associated PLD, differential e xpression of PLD isoforms, and changes in amounts of PLD protein and m RNA. Such controlled expression points to a role for PLD in membrane d eterioration and plant senescence.