SUBCELLULAR-LOCALIZATION OF ACTIN DEPOLYMERIZING FACTOR IN MATURE ANDGERMINATING POLLEN

Mature pollen contains enzymes which display unique functions during p ollen germination and tube growth. We have previously isolated pollen- specific cDNA clones from lily and Brassica napus which encode for a p utative actin depolymerizing factor (ADF). Polyclonal antibodies were raised against a synthetic peptide which corresponds to the carboxy te rminal region of the ADF proteins. Immune blot analysis showed that th e protein is preferentially expressed in the male gametophyte and is n ot detectable in other reproductive or vegetative organs, The protein was first detectable in 6.5 cm-long Easter lily buds and continued to reach a maximal level in mature flowers. Transmission electron microsc opy data revealed that the ADF was associated with electron dense incl usions which were embedded in the storage bodies of mature pollen. Dur ing a 30 min in vitro pollen germination, some of the ADF remained att ached to the inclusions. However, after 1 h incubation, most of the AD F was dissipated into the cytosol. Subcellular fractionation experimen ts confirmed the attachment of ADF with a small subcellular particle. The protein was stably maintained in cytosol during 16 h incubation, s uggesting that the ADF may play an important role during pollen tube g rowth, Our finding, along with the previous observation that the elect ron dense inclusion is a storage place for minerals and proteins, sugg ests that the inclusion may be a common location for storing nutrients and enzymes which are utilized during pollen tube growth and fertiliz ation.