Identification of a suppressor of the Dictyostelium profilin-minus phenotype as a CD36/LIMP-II homologue

Profilin is an ubiquitous G-actin binding protein in eukaryotic cells, Lack of both profilin isoforms in Dictyostelium discoideum resulted in impaired cytokinesis and an arrest in development, A restriction enzyme-mediated in tegration approach was applied to profilin-minus cells to identify suppress or mutants for the developmental phenotype, A mutant with wildtype-like dev elopment and restored cytokinesis was isolated, The gene affected was found to code for an integral membrane glycoprotein of a predicted size of 88 kD containing two transmembrane domains, one at the NH2 terminus and the othe r at the COOH terminus. It is homologous to mammalian CD36/LIMP-II and repr esents the first member of this family in D. discoideum, therefore the name DdLIMP is proposed. Targeted disruption of the lmpA gene in the profilin-m inus background also rescued the mutant phenotype. Immunofluorescence revea led a localization in vesicles and ringlike structures on the cell surface. Partially purified DdLIMP bound specifically to PIP2 in sedimentation acid gel filtration assays. A direct interaction between DdLIMP and profilin co uld not be detected, and it is unclear how far upstream in a regulatory cas cade DdLIMP might be positioned. However, the PIP2 binding of DdLIMP points towards a function via the phosphatidylinositol pathway, a major regulator of profilin.