Characterisation of Phytophthora nicotianae zoospore and cyst membrane proteins

Abstract

Motile zoospores of species of oomycetes encyst rapidly to form walled cysts that germinate and infect host plants. Differences in the protein composition of the plasma membrane and endomembranes of zoospores and cysts of the oomycete Phytophthora nicotianae have been explored by comparing patterns of polypeptides in one- and two-dimensional gels of microsomal fractions. Against a backdrop of common components, this comparison revealed that at least 53 proteins were specific to, or occurred preferentially in, the microsomal fraction of one spore type or the other. In addition, proteins common to zoospore and cyst plasma membranes were further investigated by immunocytochemical labelling with a panel of 10 monoclonal antibodies raised against P. nicotianae spore components. The results of immunolocalisation studies and immunoblotting showed that the antibodies reacted with four different sets of membrane proteins, and were grouped accordingly. Group 1 antibodies bound preferentially to the bladder of the water expulsion vacuole or a region of the cell surface associated with it. Group 2 antibodies reacted with a protein of high relative molecular weight (>200 kDa) found in zoospores and cyst plasma membranes and in the cleavage membranes of sporangia. Group 3 antibodies reacted with a set of proteins occurring in the plasma membrane of zoospores and cysts, in the peripheral cisternae, in the spongiome membranes of the water expulsion vacuole, in the cleavage membranes of sporangia, and in the plasma membrane and apical vesicle membranes in hyphae and germinating cysts. Group 4 antibodies bound to a set of proteins present in the zoospore and cyst plasma membrane, in sporangial cleavage membranes and in the spongiome but not present in the peripheral cisternae, hyphae or germinating cysts. The results document the presence of proteins that are common to the plasma membrane of all stages of the asexual life cycle of P. nicotianae, of proteins that occur in the plasma membrane of the asexual spores but not of hyphae, and of proteins that occur in the membranes of either zoospores or cysts. The results also indicate that zoospore and cyst plasma membrane proteins may be present in specific subsets of other membranes within the zoospores.