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* Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, National Institutes of
Health (NIH), Bethesda, Maryland 20892; The mechanisms of localization and retention of membrane proteins in the inner nuclear membrane and the fate of this membrane system during mitosis were studied in living cells using the inner nuclear
membrane protein, lamin B receptor, fused to green
fluorescent protein (LBR-GFP). Photobleaching techniques revealed the majority of LBR-GFP to be completely immobilized in the nuclear envelope (NE) of interphase cells, suggesting a tight binding to
heterochromatin and/or lamins. A subpopulation of
LBR-GFP within ER membranes, by contrast, was entirely mobile and diffused rapidly and freely (D = 0.41 ± 0.1 µm2/s). High resolution confocal time-lapse imaging
in mitotic cells revealed LBR-GFP redistributing into
the interconnected ER membrane system in
prometaphase, exhibiting the same high mobility and diffusion constant as observed in interphase ER membranes. LBR-GFP rapidly diffused across the cell
within the membrane network defined by the ER, suggesting the integrity of the ER was maintained in mitosis, with little or no fragmentation and vesiculation. At the end of mitosis, nuclear membrane reformation coincided with immobilization of LBR-GFP in ER elements at contact sites with chromatin. LBR-GFP-containing ER membranes then wrapped around chromatin over the course of 2-3 min, quickly and efficiently compartmentalizing nuclear material. Expansion of the NE followed over the course of 30-80 min.
Thus, selective changes in lateral mobility of LBR-GFP
within the ER/NE membrane system form the basis for
its localization to the inner nuclear membrane during interphase. Such changes, rather than vesiculation
mechanisms, also underlie the redistribution of this
molecule during NE disassembly and reformation in
mitosis.
Department of Physics, Cornell University, Ithaca, New York 14853; § Light Imaging
Facility, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, Maryland 20892; and Departments of
Medicine and of Anatomy and Cell Biology, College of Physicians and Surgeons, Columbia University, New York 10032
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