Tracking the interactions of rRNA processing proteins during nucleolar assembly in living cells.
Reorganization of the nuclear machinery after mitosis is a fundamental but poorly understood process. Here, we investigate the recruitment of the nucleolar processing proteins in the nucleolus of living cells at the time of nucleus formation. We question the role of the prenucleolar bodies (PNBs), during migration of the processing proteins from the chromosome periphery to sites of rDNA transcription. Surprisingly, early and late processing proteins pass through the same PNBs as demonstrated by rapid two-color four-dimensional imaging and quantification, whereas a different order of processing protein recruitment into nucleoli is supported by differential sorting. Protein interactions along the recruitment pathway were investigated using a promising time-lapse analysis of fluorescence resonance energy transfer. For the first time, it was possible to detect in living cells the interactions between proteins of the same rRNA processing machinery in nucleoli. Interestingly interactions between such proteins also occur in PNBs but not at the chromosome periphery. The dynamics of these interactions suggests that PNBs are preassembly platforms for rRNA processing complexes.
Angelier
Nicole
N
Nuclei and Cell Cycle Laboratory, Institut Jacques Monod, Centre National de la Recherche Scientifique, University Paris VI and Paris VII, 75251 Paris, France.
Tramier
Marc
M
Louvet
Emilie
E
Coppey-Moisan
Maïté
M
Savino
Tula M
TM
De Mey
Jan R
JR
Hernandez-Verdun
Danièle
D
D000705:Anaphase; D002461:Cell Line, Transformed; D002466:Cell Nucleolus / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D016193:G1 Phase; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007700:Kinetics; D018715:Microscopy, Video; D008938:Mitosis; D008954:Models, Biological; D009687:Nuclear Proteins / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012335:RNA, Ribosomal / Q000378:metabolism*; D013692:Telophase; D014158:Transcription, Genetic
D002868:Chromosomal Proteins, Non-Histone; D009687:Nuclear Proteins; D012335:RNA, Ribosomal; C107468:RRP1 protein, human; C047458:fibrillarin; D049452:Green Fluorescent Proteins