A cell number counting factor regulates Akt/protein kinase B to regulate Dictyostelium discoideum group size.
Little is known about how individual cells can organize themselves to form structures of a given size. During development, Dictyostelium discoideum aggregates in dendritic streams and forms groups of approximately 20,000 cells. D. discoideum regulates group size by secreting and simultaneously sensing a multiprotein complex called counting factor (CF). If there are too many cells in a stream, the associated high concentration of CF will decrease cell-cell adhesion and increase cell motility, causing aggregation streams to break up. The pulses of cyclic AMP (cAMP) that mediate aggregation cause a transient translocation of Akt/protein kinase B (Akt/PKB) to the leading edge of the plasma membrane and a concomitant activation of the kinase activity, which in turn stimulates motility. We found that countin- cells (which lack bioactive CF) and wild-type cells starved in the presence of anticountin antibodies (which block CF activity) showed a decreased level of cAMP-stimulated Akt/PKB membrane translocation and kinase activity compared to parental wild-type cells. Recombinant countin has the bioactivity of CF, and a 1-min treatment of cells with recombinant countin potentiated Akt/PKB translocation to membranes and Akt/PKB activity. Western blotting of total cell lysates indicated that countin does not affect the total level of Akt/PKB. Fluorescence microscopy of cells expressing an Akt/PKB pleckstrin homology domain-green fluorescent protein (PH-GFP) fusion protein indicated that recombinant countin and anti-countin antibodies do not obviously alter the distribution of Akt/PKB PH-GFP when it translocates to the membrane. Our data indicate that CF increases motility by potentiating the cAMP-stimulated activation and translocation of Akt/PKB.
Department of Biochemistry and Cell Biology, MS-140, Rice University, 6100 S. Main St., Houston, TX 77005-1892, USA.
Tong
T
Gao
David
D
Knecht
Lei
L
Tang
R Diane
RD
Hatton
Richard H
RH
Gomer
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D004023:Dictyostelium / Q000166:cytology* / Q000201:enzymology* / Q000235:genetics; D017125:Genes, Protozoan; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D011518:Proto-Oncogene Proteins; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C120918:countin protein, Dictyostelium discoideum; D049452:Green Fluorescent Proteins; D000242:Cyclic AMP; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt