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Related Experiment Videos

Mitotic chromosomal bcl-2. II. Localization to interphase nuclei

C A Schandl1, S Li, G G Re

  • 1Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, South Carolina, USA.

The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society
|January 16, 1999
PubMed
Summary

We discovered that bcl-2 protein is located in the nucleus during interphase, not just on mitotic chromosomes. This nuclear localization may be due to epitope masking or conformational changes, requiring further investigation.

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Area of Science:

  • Molecular Biology
  • Cell Biology
  • Oncology

Background:

  • Previous studies indicated bcl-2 protein is exclusively in mitotic chromosomes using immunofluorescence.
  • Conflicting Western blot analyses suggested bcl-2 levels remain constant throughout the cell cycle.
  • The intracellular localization of bcl-2 during interphase remained unclear.

Purpose of the Study:

  • To investigate the distribution of bcl-2 protein during the interphase stage of the cell cycle.
  • To reconcile conflicting data regarding bcl-2 localization and cell cycle progression.
  • To identify the cellular compartment containing bcl-2 during interphase.

Main Methods:

  • Immunofluorescence microscopy on fixed cells.
  • Biochemical extraction and immunoprecipitation.

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  • Western blotting with specific antibodies (anti-bcl-2 and anti-Bax).
  • Classical cell fractionation to isolate nuclei, cytosol, and cell membranes.

    • Main Results:

    • Bcl-2 protein was detected in the nucleus during early G2 phase by immunofluorescence.
    • Immunoprecipitation confirmed the presence of bcl-2 in interphase cells, despite lack of immunofluorescence signal.
    • Cell fractionation revealed that the majority of bcl-2 resides in the nucleus during interphase.
    • Interphase nuclear bcl-2 was not found to be complexed with Bax, unlike in overexpressing cells.

    Conclusions:

    • The lack of immunofluorescence signal for bcl-2 in interphase cells is likely due to epitope masking or conformational changes within the nucleus.
    • Nuclear localization of bcl-2 during interphase represents a novel finding.
    • Further research is needed to elucidate the function of this nuclear bcl-2 form and its potential relationship with pre-mitotic phosphorylation.