Live fluorescence imaging reveals early recruitment of emerin, LBR, RanBP2, and Nup153 to reforming functional nuclear envelopes.
We determined the times when the nuclear membrane, nuclear pore complex (NPC) components, and nuclear import function were recovered during telophase in living HeLa cells. Simultaneous observation of fluorescently-labeled NLS-bearing proteins, lamin B receptor (LBR)-GFP, and Hoechst33342-stained chromosomes revealed that nuclear membranes reassembled around chromosomes by 5 minutes after the onset of anaphase (early telophase) whereas nuclear import function was recovered later, at 8 minutes. GFP-tagged emerin also accumulated on chromosomes 5 minutes after the onset of anaphase. Interestingly, emerin and LBR initially accumulated at distinct, separate locations, but then became uniform 8 minutes after the onset of anaphase, concurrent with the recovery of nuclear import function. We further determined the timing of NPC assembly by immunofluorescence staining of cells fixed at precise times after the onset of anaphase. Taken together, these results showed that emerin, LBR, and several NPC components (RanBP2, Nup153, p62), but not Tpr, reconstitute around chromosomes very early in telophase prior to the recovery of nuclear import activity.
Haraguchi
T
T
Kansai Advanced Research Center, Communications Research Laboratory, CREST Research Project, Japan Science and Technology Corporation, Iwaoka-cho, Nishi-ku, Kobe 651-2492, Japan. tokuko@crl.go.jp
Koujin
T
T
Hayakawa
T
T
Kaneda
T
T
Tsutsumi
C
C
Imamoto
N
N
Akazawa
C
C
Sukegawa
J
J
Yoneda
Y
Y
Hiraoka
Y
Y
D001692:Biological Transport / Q000502:physiology; D004268:DNA-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D006367:HeLa Cells / Q000378:metabolism / Q000502:physiology; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism / Q000502:physiology*; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones; D009685:Nuclear Envelope / Q000378:metabolism / Q000502:physiology*; D019913:Nuclear Localization Signals / Q000502:physiology; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins / Q000378:metabolism / Q000502:physiology*; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism / Q000502:physiology*; D013692:Telophase / Q000502:physiology; D013946:Thymopoietins / Q000378:metabolism / Q000502:physiology*; D000097682:Lamin B Receptor
D004268:DNA-Binding Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D019913:Nuclear Localization Signals; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D013946:Thymopoietins; D000097682:Lamin B Receptor; C079377:NUP153 protein, human; C092710:emerin; C094493:ran-binding protein 2