Subcellular localization of the retinoblastoma protein.

L. Szekely, E. Uzvolgyi, W. Q. Jiang, M. Durko, K. G. Wiman, G. Klein, J. Sumegi

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39 Scopus citations

Abstract

The subcellular localization of the retinoblastoma (RB) protein has been studied in primate cell lines by immunofluorescence staining using different monoclonal and polyclonal antibodies. The protein appeared as granules of heterogeneous size over the interphase nuclei. Computer assisted digital overlap analysis indicated that it was predominantly localized in euchromatic areas with low DNA density. The largest RB positive grains lined up on the heterochromatin/euchromatin boundary. During mitosis, the RB protein dissociated from the condensing chromosomes. It was dispersed throughout the cytoplasm during metaphase and anaphase, and it reassociated with the decondensing chromatin during telophase. A new monoclonal antibody, designated aRB1C1, was raised against a bacterial TrpE/human retinoblastoma protein. It specifically recognized the nonphosphorylated and differentially phosphorylated forms of the RB protein in immunoprecipitation experiments. A collection of RB expressing cell lines gave a positive staining reaction with the antibody, whereas the RB negative Weri-RB-27 retinoblastoma and OHS osteosarcoma cells failed to react. Wild-type RB complementary DNA was introduced into Weri-RB-27 by retrovirus mediated gene transfer. Similar experiments were performed with the DU145 prostatic carcinoma cell line that expresses a mutant RB protein. Reconstituted cells of both lines expressed the normal size RB protein and gave a positive immunofluorescence reaction with the aRB1C1 and other anti-RB antibodies. The new monoclonal antibody, however, showed cell type dependent differences of the staining pattern compared to other anti-RB antibodies, suggesting differentiation dependent accessibility to its epitope.

Original languageEnglish (US)
Pages (from-to)287-295
Number of pages9
JournalCell growth & differentiation : the molecular biology journal of the American Association for Cancer Research
Volume2
Issue number6
StatePublished - Jun 1991

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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    Szekely, L., Uzvolgyi, E., Jiang, W. Q., Durko, M., Wiman, K. G., Klein, G., & Sumegi, J. (1991). Subcellular localization of the retinoblastoma protein. Cell growth & differentiation : the molecular biology journal of the American Association for Cancer Research, 2(6), 287-295.