PRC1 components exhibit different binding kinetics in Polycomb bodies.
BACKGROUND
Polycomb group (PcG) proteins keep the memory of cell identity by maintaining the repression of numerous target genes. They accumulate into nuclear foci called Polycomb bodies, which function in Drosophila cells as silencing compartments where PcG target genes convene. PcG proteins also exert their activities elsewhere in the nucleoplasm. In mammalian cells, the dynamic organisation and function of Polycomb bodies remain to be determined.


RESULTS
Fluorescently tagged PcG proteins CBXs and their partners BMI1 and RING1 form foci of different sizes and intensities in human U2OS cells. Fluorescence recovery after photobleaching (FRAP) analysis reveals that PcG dynamics outside foci is governed by diffusion as complexes and transient binding. In sharp contrast, recovery curves inside foci are substantially slower and exhibit large variability. The slow binding component amplitudes correlate with the intensities and sizes of these foci, suggesting that foci contained varying numbers of binding sites. CBX4-green fluorescent protein (GFP) foci were more stable than CBX8-GFP foci; yet the presence of CBX4 or CBX8-GFP in the same focus had a minor impact on BMI1 and RING1 recovery kinetics.


CONCLUSIONS
We propose that FRAP recovery curves provide information about PcG binding to their target genes outside foci and about PcG spreading onto chromatin inside foci.
Biophotonique Cellulaire Fonctionnelle, Interdisciplinary Research Institute, Université Lille 1 - CNRS USR 3078, Parc de la Haute Borne, Villeneuve d'Ascq, 59658, France.
Bernard
B
Vandenbunder
Nicolas
N
Fourré
Aymeric
A
Leray
Florian
F
Mueller
Pamela
P
Völkel
Pierre-Olivier
PO
Angrand
Laurent
L
Héliot
D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D045744:Cell Line, Tumor; D006801:Humans; D007700:Kinetics; D063146:Polycomb-Group Proteins / Q000737:chemistry* / Q000378:metabolism*
D018797:Cell Cycle Proteins; C116662:PRC1 protein, human; D063146:Polycomb-Group Proteins
Binding kinetics; Diffusion; FRAP; Polycomb bodies; Transcriptional regulation