First published online 27 November 2002
doi: 10.1242/jcs.00225
Polycomb group gene silencing proteins are concentrated in the perichromatin compartment of the mammalian nucleus
Dusan Cmarko1,
Pernette J. Verschure2,
Arie P. Otte2,
Roel van Driel2 and
Stanislav Fakan1,*
1 Centre of Electron Microscopy, University of Lausanne, 27 Bugnon, CH-1005
Lausanne, Switzerland
2 Swammerdam Institute for Life Sciences, BioCentrum Amsterdam, University of
Amsterdam, P.O. Box 94062, 1090 GB Amsterdam, The Netherlands
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Fig. 1. Immunofluorescent labelling of the PcG proteins in cultured cells. Confocal
optical section through the central part of the nucleus. (A-E) SW480 cells
labelled with anti-BMI1 (A), anti-HPC2 (B) and DAPI (C); (D) shows the merged
image and (E) a line scan along the white arrow in panel D, showing the
distribution of BMI1 (red line), HPC2 (green line) and DAPI (blue line). (F-G)
SW 480 cells labelled with anti-RING1 (F) and DAPI (G). (H-J) T24 cells
labelled with anti-BMI1 (H) and DAPI (I); J shows a line scan along the white
arrow in H, showing the distribution of BMI1 (green line) and DAPI (blue
line). (K-M) T24 cells labelled with anti-HPH1 (K) and DAPI (L); (M) shows a
line scan along the white arrow in K, showing the distribution of HPH1 (green
line) and DAPI (blue line).
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Fig. 2. Immunoelectron microscopic visualisation of PcG protein distribution in
sections of paraformaldehyde-fixed SW480 cells, following the EDTA staining
procedure. (A) Rabbit anti-HPH1 antibody revealed by goat anti-rabbit
secondary antibody conjugated with 12 nm colloidal gold particles; (B) rabbit
anti-HPC2 antibody and 12 nm particles; (C) rabbit anti-RING1 and 12 nm
particles and (D) rabbit anti-BMI1 and 15 nm particles. Nuclear distribution
of PcG proteins frequently occurs on electron-dense RNP-containing fibrils,
which are distributed in the nucleoplasm (some fibrils indicated by
arrowheads). Condensed chromatin (c) is virtually devoid of signal. Bars, 0.3
µm.
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Fig. 3. Immunoelectron microscopic visualisation of PcG proteins in (A) an
osmium-ammine-stained section of a paraformaldehyde-fixed SW480 cell
(anti-HPC2 labelling) and (B) in high pressure frozen and cryosubstituted rat
liver tissue (anti-BMI labelling); uranyl-lead stained. The primary antibodies
were revealed by goat anti-rabbit secondary antibody conjugated with 12 nm
colloidal gold particles. PcG proteins mainly occur at the periphery of
condensed chromatin (c). Some perichromatin fibrils are indicated by
arrowheads. Bars, 0.2 µm.
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Fig. 4. Double immunolabelling. (A) Double-labelling with mouse monoclonal
anti-HPC2 antibody (15 nm gold particles) and rabbit anti-BMI1 antibody (10 nm
gold particles, some indicated by arrowheads) in an SW480 cell stained with
osmium ammine specific for DNA; most signal is observed on the periphery or
outside DNA-rich regions. (B) Double labelling experiment with PcG protein
BMI1 (6 nm gold particles) and nascent BrU-containing RNA (15 nm particles) in
a T24 cell; EDTA staining. Both labels mostly occur near the border of
condensed chromatin regions (c), sometimes in association with perichromatin
fibrils (some indicated by arrowheads). A cluster of interchromatin granules
(large arrow) exhibits anti-PcG protein label only on its periphery (small
arrows). Bars, 0.2 µm.
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© The Company of Biologists Ltd 2003
