The nuclear envelope (NE) is a double membrane that segregates nuclear components from the cytoplasm in eukaryotic cells. It is well-known that the NE undergoes a breakdown and reformation during mitosis in animal cells. However, the detailed mechanisms of the NE dynamics are not yet fully understood. Here, we propose a method for the fluorescent labeling of the NE in living cells, which enables the tracing of the NE dynamics during cell division under physiological conditions. In our method, labeling of the NE is accomplished by fixing green fluorescent protein carrying the nuclear localization signal on the inner nuclear membrane based on a unique biotinylation reaction from the archaeon Sulfolobus tokodaii. With this method, we observed HeLa cells during mitosis by confocal laser scanning microscopy and succeeded in clearly visualizing the difference in the timing of the formation of the NE and the nuclear lamina.

During mitosis in metazoan species, the nuclear envelope (NE) undergoes breakdown, and its fragments are absorbed within the membranous network of the endoplasmic reticulum (ER). Past observations by fluorescence microscopy led researchers to think that the NE loses its identity when it is absorbed within the ER membrane. However, in our previous work, we developed a more specific labelling method and found evidence that the NE does not completely lose its identity during mitosis. In the present work, we conduct further experiments, the results of which support the idea that the NE partially retains its identity during mitosis.

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