Alterations in immunolocalization of the phosphoprotein B23 in HeLa cells during serum starvation.
Bright nucleolar immunofluorescence was observed in HeLa S3 cells by immunostaining with a monoclonal antibody to the nucleolar phosphoprotein B23 (MW 37 kD/pI 5.1). After 48 h of incubation in a serum-free medium, the nucleolar fluorescence was diminished and a general nuclear immunofluorescence was observed. This change in localization of fluorescence indicated that protein B23 had migrated out of the nucleoli. No gross morphological change in nucleoli was observed by light microscopy and the immunolocalization of another nucleolar phosphoprotein, C23, was unaffected by serum deprivation. Relocation of protein B23 in nucleoli was observed after refeeding with serum-containing medium. This re-entry process was not observed after treatment with actinomycin D (50 ng/ml-5 micrograms/ml), but the process was unaffected by cycloheximide (0.2 mM). Quantitation of protein B23 in the nucleoli of the control (fed) or starved HeLa cells was done by ELISA immunoassay. A marked decrease in the amount of protein B23 occurred in the nucleoli of the starved cells (11.8 micrograms B23/mgDNA) as compared with the control nucleoli (20.8 micrograms B23/mgDNA). The amount of protein B23 in the nucleoplasm (excluding nucleoli) was 70% higher in the starved cells. Protein B23 was analysed by one- and two-dimensional PAGE. Three components of protein B23 with slightly different molecular weights and pIs (37 kD/5.1, 35 kD/5.1 and 35 kD/5.3) were observed in nucleoli. The lower molecular weight components were predominantly found in the nucleoplasm.
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D001769:Blood; D002455:Cell Division; D002466:Cell Nucleolus / Q000032:analysis* / Q000648:ultrastructure; D002467:Cell Nucleus / Q000032:analysis*; D003470:Culture Media; D003513:Cycloheximide / Q000494:pharmacology; D004247:DNA / Q000032:analysis; D003609:Dactinomycin / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D008970:Molecular Weight; D009687:Nuclear Proteins; D000090243:Nucleophosmin; D010750:Phosphoproteins / Q000032:analysis; D014176:Protein Biosynthesis; D012313:RNA / Q000096:biosynthesis; D012261:Ribonucleoproteins / Q000032:analysis* / Q000302:isolation & purification
D003470:Culture Media; C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D010750:Phosphoproteins; D012261:Ribonucleoproteins; C036825:phosphoprotein C23; D000090243:Nucleophosmin; D003609:Dactinomycin; D012313:RNA; D004247:DNA; D003513:Cycloheximide