Retinoylation of vimentin in the human myeloid leukemia cell line HL60.
Retinoylation (retinoic acid acylation) is a posttranslational modification of proteins occurring in many eukaryotic cell lines. The widespread occurrence of retinoylation suggests that it may play a role in many effects of retinoic acid (RA) on cells. The regulatory subunits of cyclic AMP-dependent protein kinase are retinoylated in the human myeloid leukemia cell line HL60 (Takahashi, N., Liapi, C., Anderson, W. B., and Breitman, T. R. (1991) Arch. Biochem. Biophys. 290, 293-302), and cytokeratins are retinoylated in normal human keratinocytes (Takahashi, N., Jetten, A. M., and Breitman, T. R. (1991) Biochem. Biophys. Res. Commun. 180, 393-400). We show, in this study, that the intermediate filament protein vimentin is retinoylated in HL60 cells during a 24-h exposure to 100 nM [3H]RA. We found that a retinoylated protein of M(r) 55,000 coeluted on anion exchange chromatography and comigrated on either one- or two-dimensional polyacrylamide gel electrophoresis with a protein that also was stained on immunoblots by an anti-vimentin antibody. About 50% of the [3H]RA was released from this M(r) 55,000 retinoylated protein after hydrolysis with either NH2OH (1 M, pH 10) or CH3OH, 0.1 M KOH. These results indicated that a large fraction of the RA was bound to vimentin by an ester bond. Both the M(r) 55,000 retinoylated protein and immunoreactive vimentin were associated with cell nuclei isolated by two procedures. They were detached during exposure to a nonionic detergent buffer, suggesting that they are bound to the nuclear envelope. These results indicate that retinoylation is a new modification of vimentin that may be an early event in RA-induced differentiation of HL60 cells.
Takahashi
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Laboratory of Biological Chemistry, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
Breitman
T R
TR
D002454:Cell Differentiation; D002852:Chromatography, Ion Exchange; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007951:Leukemia, Myeloid; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism; D012394:Rosaniline Dyes; D013194:Staining and Labeling; D014212:Tretinoin / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D014746:Vimentin / Q000302:isolation & purification / Q000378:metabolism*
D011506:Proteins; D012394:Rosaniline Dyes; D014746:Vimentin; D014212:Tretinoin; C048139:Coomassie blue