EXPRESSION AND CHARACTERIZATION OF A DICTYOSTELIUM-DISCOIDEUM ANNEXIN

The annexins are calcium-dependent phospholipid-binding proteins. Rece ntly the gene encoding the homologue of a mammalian annexin has been i dentified in Dictyostelium discoideum. Analysis of cDNA and genomic cl ones showed that the transcript for Dictyostelium annexin is alternati vely spliced (Greenwood, M. and Tsang, A. (1991) Biochim. Biophys. Act a 1088, 429-432; Doring, V., Schleicher, M and Noegel, A. (1991) J. Bi ol. Chem. 266, 17509-17515). Here, we showed that the Dictyostelium an nexin DNA hybridized to two populations of transcripts. We used a reco mbinant annexin polypeptide to raise polyclonal antibody. Immunoblot a nalysis revealed that the antibody recognized two polypeptides of 48 k Da and 54 kDa in developing D. discoideum cells. The molecular sizes o f these polypeptides correspond well with the expected sizes of the al ternatively spliced products. The 48-kDa and 54-kDa polypeptides were purified by isoelectric focusing to more than 70% homogeneity. The par tially purified proteins were found to associate with phosphatidylseri ne vesicles in a calcium-dependent manner. These results suggest that the 48- and 54-kDa polypeptides are the products of alternative splici ng of the annexin transcripts. During development the two polypeptides accumulate at different rates to about 60 times the level detected in vegetative cells. On the other hand, RNA blot analysis showed that th e level of the annexin transcripts in multicellular aggregates was abo ut 5 times that of vegetative cells.