Live imaging system for visualizing nuclear pore complex (NPC) formation during interphase in mammalian cells.
Nuclear pore complexes (NPCs) are 'supramolecular complexes' on the nuclear envelope assembled from multiple copies of approximately 30 different proteins called nucleoporins (Nups) that provide aqueous channels for nucleocytoplasmic transport during interphase. Although the structural aspects of NPCs have been characterized in detail, NPC formation and its regulation, especially during interphase, are poorly understood. In this study, using the temperature-sensitive RCC1 mutant tsBN2, a baby hamster kidney 21 cell line, we found that a lack of RCC1 activity inhibited NPC formation during interphase, suggesting that RanGTP is required for NPC formation during interphase in mammalian cells. Utilizing the reversible RCC1 activity in tsBN2 cells, we established a live-cell system that allows for the inhibition or initiation of NPC formation by changes in temperature. Our system enables the examination of NPC formation during interphase in living cells. As a lack of RCC1 decreased some Nups containing unstructured phenylalanine-glycine repeats in the NPC structure, we propose that RCC1 is also involved in maintaining NPC integrity during interphase in mammalian cells.
Iino
Haruki
H
Cellular Dynamics Laboratory, RIKEN Advanced Science Institute, Wako, Saitama 351-0198, Japan.
Maeshima
Kazuhiro
K
Nakatomi
Reiko
R
Kose
Shingo
S
Hashikawa
Tsutomu
T
Tachibana
Taro
T
Imamoto
Naoko
N
D021581:Active Transport, Cell Nucleus / Q000502:physiology; D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D007399:Interphase / Q000502:physiology*; D008855:Microscopy, Electron, Scanning; D009154:Mutation; D022022:Nuclear Pore / Q000378:metabolism* / Q000648:ultrastructure; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D038761:Photobleaching; D013696:Temperature; D013997:Time Factors
D018797:Cell Cycle Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; D049452:Green Fluorescent Proteins