A Dictystelium mutant with defective aggregate size determination.
Starved Dictyostelium cells aggregate into groups of roughly 10(5) cells. We have identified a gene which, when repressed by antisense transformation or homologous recombination, causes starved cells to form large numbers of small aggregates. We call the gene smlA for small aggregates. A roughly 1.0 kb smlA mRNA is expressed in vegetative and early developing cells, and the mRNA level then decreases at about 10 hours of development. The sequence of the cDNA and the derived amino acid sequence of the SmlA protein show no significant similarity to any known sequence. There are no obvious motifs in the protein or large regions of hydrophobicity or charge. Immunofluorescence and staining of Western blots of cell fractions indicates that SmlA is a 35x10(3) Mr cytosolic protein present in all vegetative and developing cells and is absent from smlA cells. The absence of SmlA does not affect the growth rate, cell cycle, motility, differentiation, or developmental speed of cells. Synergy experiments indicate that mixing 5% smlA cells with wild-type cells will cause the wild-type cells to form smaller fruiting bodies and aggregates. Although there is no detectable SmlA protein secreted from cells, starvation medium conditioned by smlA cells will cause wild-type cells to form large numbers of small aggregates. The component in the smlA-conditioned media that affects aggregate size is a molecule with a molecular mass greater than 100x10(3) Mr that is not conditioned media factor, phosphodiesterase or the phosphodiesterase inhibitor. The data thus suggest that the cytosolic protein SmlA regulates the secretion or processing of a secreted factor that regulates aggregate size.
Brock
D A
DA
Howard Hughes Medical Institute, Department of Biochemistry and Cell Biology, Rice University, Houston, TX 77251-1892, USA.
Buczynski
G
G
Spann
T P
TP
Wood
S A
SA
Cardelli
J
J
Gomer
R H
RH
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000502:physiology; D002453:Cell Cycle; D017077:Culture Media, Conditioned; D000242:Cyclic AMP / Q000502:physiology; D016373:DNA, Antisense / Q000235:genetics; D003911:Dextrans / Q000378:metabolism; D004023:Dictyostelium / Q000235:genetics / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D005089:Exocytosis; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015966:Gene Expression Regulation, Fungal; D008969:Molecular Sequence Data; D009068:Movement; D009154:Mutation; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D014170:Transformation, Genetic
D015815:Cell Adhesion Molecules; D017077:Culture Media, Conditioned; D016373:DNA, Antisense; D003911:Dextrans; D005656:Fungal Proteins; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C101537:SmlA protein, Dictyostelium discoideum; C052291:cell cohesion molecule, Dictyostelium; C015219:fluorescein isothiocyanate dextran; D000242:Cyclic AMP; D010727:Phosphoric Diester Hydrolases; D016650:Fluorescein-5-isothiocyanate