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 integration approach was applied to profilin-minus cells to identify suppressor mutants for the developmental phenotype. A mutant with wild-type-like development 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 other at the COOH terminus. It is homologous to mammalian CD36/LIMP-II and represents the first member of this family in D. discoideum, therefore the name DdLIMP is proposed. Targeted disruption of the lmpA gene in the profilin-minus background also rescued the mutant phenotype. Immunofluorescence revealed a localization in vesicles and ringlike structures on the cell surface. Partially purified DdLIMP bound specifically to PIP2 in sedimentation and gel filtration assays. A direct interaction between DdLIMP and profilin could not be detected, and it is unclear how far upstream in a regulatory cascade DdLIMP might be positioned. However, the PIP2 binding of DdLIMP points towards a function via the phosphatidylinositol pathway, a major regulator of profilin.
Karakesisoglou
I
I
A.-Butenandt-Institut für Zellbiologie, Ludwig-Maximilians-Universität, 80336 München, Germany.
Janssen
K P
KP
Eichinger
L
L
Noegel
A A
AA
Schleicher
M
M
D000595:Amino Acid Sequence; D000818:Animals; D018955:CD36 Antigens / Q000737:chemistry; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D016384:Consensus Sequence; D003285:Contractile Proteins; D004023:Dictyostelium / Q000235:genetics*; D005786:Gene Expression Regulation; D018390:Gene Targeting; D017125:Genes, Protozoan; D016153:Genes, Suppressor; D008247:Lysosomes / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000502:physiology; D008840:Microfilament Proteins / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D010641:Phenotype; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D012150:Polymorphism, Restriction Fragment Length; D051304:Profilins; D015800:Protozoan Proteins / Q000235:genetics* / Q000502:physiology; D011995:Recombination, Genetic; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
D018955:CD36 Antigens; D003285:Contractile Proteins; D008562:Membrane Glycoproteins; D008840:Microfilament Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D051304:Profilins; D015800:Protozoan Proteins