Cytoskeletal association of an esterase in Dictyostelium discoideum.
A 70-kDa glycoprotein, gp70, was found enriched in the detergent-insoluble cytoskeletal fraction of axenically grown Dictyostelium discoideum cells. Its N-terminal amino acid sequence identified it as 'crystal protein' (L. Bomblies et al., 1990, J. Cell Biol. 110, 669-679). This finding was corroborated when antibody to crystal protein cross-reacted with gp70 and its deglycosylated form. The postulated esterase activity of gp70/crystal protein was verified through comparative enzyme assays of extracts derived from cells that either overexpressed or lacked gp70. Gp70 cosedimented with cytoskeletons on sucrose gradients, suggesting an interaction with the cytoskeleton. Coisolation of gp70 with detergent-extracted cells, observed by immunofluorescence microscopy, also implied a gp70-cytoskeletal association. These data supported the idea that the localization or secretion of gp70, or both, was cytoskeletally mediated. Although axenically grown cells contained high levels of gp70, the same cell lines had reduced levels of gp70 when grown in bacterial suspension or in nutrient media containing bacteria. Bacterially grown cells, compared to axenically grown cells, had lower fluid-phase uptake rates even when nutrient media was present, indicating that phagocytosis was a preferred mode of feeding. Thus, bacteria inhibited gp70 expression, which suggested a role for prestarvation factor, in regulating its synthesis.
School of Biological Sciences, University of Nebraska-Lincoln, Lincoln, Nebraska, 68588-0118, USA. cpc@unlinfo.unl.edu
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D000595:Amino Acid Sequence; D000818:Animals; D003599:Cytoskeleton / Q000201:enzymology* / Q000378:metabolism; D004023:Dictyostelium / Q000166:cytology / Q000201:enzymology* / Q000254:growth & development; D004789:Enzyme Activation; D004950:Esterases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D015800:Protozoan Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
D006023:Glycoproteins; D015800:Protozoan Proteins; C063332:crystal protein, Dictyostelium; D004950:Esterases