A ras-encoded protein in Dictyostelium discoideum is acylated and membrane-associated.
Dictyostelium discoideum synthesizes a 23,000 Mr protein, p23dd-ras, closely related to the mammalian oncogene-encoded protein p21ras. To investigate the subcellular localization of p23dd-ras, conditions were optimized to reduce protein degradation following cell breakage. Subcellular fractionation of D. discoideum showed that p23dd-ras was associated predominantly with the membrane fraction during both vegetative growth and differentiation. In the absence of suitable protease inhibitors considerable amounts of a truncated form of p23dd-ras were recovered in the cytosol fraction, suggesting that intact p23dd-ras is attached to the membrane by a short terminal peptide sequence. Radio-isotope labelling of D. discoideum with myristic acid or palmitic acid in the presence of excess unlabelled acetate resulted in radio-isotope incorporation into a select group of proteins including p23dd-ras. No acyl label appeared in the truncated cytoplasmic form of p23dd-ras when cell breakage was performed in the absence of suitable protease inhibitors, indicating that the acyl group is associated with the short terminal peptide that is cleaved. These data suggest that p23dd-ras, like its mammalian counterpart, is acylated and associated with the plasma membrane. There was no evidence during a 30-minute pulse of methionine for a cytoplasmic precursor to the membrane-bound p23dd-ras, suggesting that the turnover of the presumptive precursor must be much more rapid in D. discoideum than for pro-p21ras in mammalian cells.
Weeks
G
G
Department of Microbiology, University of British Columbia, Vancouver, Canada.
Lima
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AF
Pawson
T
T
D000085:Acetates / Q000378:metabolism; D019342:Acetic Acid; D000215:Acylation; D002462:Cell Membrane / Q000378:metabolism; D004023:Dictyostelium / Q000235:genetics* / Q000378:metabolism; D005656:Fungal Proteins / Q000096:biosynthesis / Q000235:genetics*; D008715:Methionine / Q000378:metabolism; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D013462:Sulfur Radioisotopes; D014316:Tritium; D018631:ras Proteins
D000085:Acetates; D005656:Fungal Proteins; D009227:Myristic Acids; D013462:Sulfur Radioisotopes; C489016:ras protein, Dictyostelium; D019814:Myristic Acid; D014316:Tritium; D008715:Methionine; D018631:ras Proteins; D019342:Acetic Acid