Lamin B receptor (LBR), a chromatin and lamin B-binding protein in the inner nuclear membrane, has been proposed to target the membrane precursor vesicles to chromatin mediated by importin ␤ during the nuclear envelope (NE) assembly. However, the mechanisms for the binding of LBR with importin ␤ and the membrane targeting by LBR in NE assembly remain largely unknown. In this report, we show that the amino acids (aa) 69 -90 of LBR sequences are required to bind with importin ␤ at aa 45-462, and the binding is essential for the NE membrane precursor vesicle targeting to the chromatin during the NE assembly at the end of mitosis. We also show that this binding is cell cycle-regulated and dependent on the phosphorylation of LBR Ser-71 by p34 cdc2 kinase. RNAi knockdown of LBR causes the NE assembly failure and abnormal chromatin decondensation of the daughter cell nuclei, leading to the daughter cell death at early G 1 phase by apoptosis. Perturbation of the interaction of LBR with importin ␤ by deleting the LBR N-terminal spanning region or aa 69 -73 also induces the NE assembly failure, the abnormal chromatin decondensation, and the daughter cell death. The first transmembrane domain of LBR promotes the NE production and expansion, because overexpressing this domain is sufficient to induce membrane overproduction of the NE. Thus, these results demonstrate that LBR targets the membrane precursor vesicles to chromatin by interacting with importin ␤ in a LBR phosphorylation-dependent manner during the NE assembly at the end of mitosis and that the first transmembrane domain of LBR promotes the LBR-bearing membrane production and the NE expansion in interphase.Nucleus, the largest organelle that contains the genome of the eukaryotic organisms, is surrounded by a continuous nuclear envelope (NE) 2 composed of a pair of inner and outer nuclear membranes, studded by numerous nuclear pore complexes. Through the nuclear pore complexes, the NE controls the flow of molecules between the nucleus and the cytoplasm in a tightly regulated manner (1-5). The NE is highly dynamic during the cell cycle. It disassembles into membranous vesicles or tubules and disperses into the cytoplasm at the onset of prometaphase and reassembles around the newly replicated chromosomes at the end of mitosis using their previously disassembled components (2, 4, 6). Recently, it was shown that Ran, a small Ras-like nuclear GTPase, and its binding proteins regulate the NE assembly process (7-13). Ran exists in GTP-and GDP-bound states that interact differently with effectors. Conversion between these two states and the assembly or disassembly of the effector complexes requires the interaction of regulator proteins, the nucleus-based guanine nucleotide exchange factor RCC1 and the cytoplasm-localized GTPase-activating protein RanGAP1 (6,14). The compartmentalization of the regulators produces a high concentration gradient of Ran-GTP in the nucleus and Ran-GDP in the cytoplasm. In mitosis, after the NE breakdown, the nucleoplasm and the cytoplasm are ...

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