Direct contribution of ER cisternae to assembly of the nuclear envelope. (A and B) 3D renderings and serial optical sections show the relationship between the ER cisternae and nascent nuclear envelope in HeLa (A) and BSC1 (B) cells. 3D image stacks were acquired from mitotic cells expressing GFP-Sec61β and H2B-RFP. (a) Fluorescent image from the middle section. The boxed region contains an example of ER cisternae in continuity with the nascent nuclear envelope and is further shown in b together with evenly spaced serial images along the z axis. The section at 0.00 µm corresponds to the optical section in a. b′ displays rendered 3D views of region b. The example highlights a direct contact between ER cisternae (green lines) and the nascent nuclear envelope (NE; white lines). (C) EM tomograms of the nascent nuclear envelope from an anaphase BSC1 cell. (a) Low magnification image of the whole cell. (b and c) A representative EM tomographic slice from the boxed region is shown without (b) and with (c) superimposition of a model for ER cisterna (green) and ribosomes (magenta). Chrom., chromosome mass. (d–f) Three views of the rendered 3D model. ER membrane, ribosome, and the surface of the chromosome mass are colored in green, magenta, and blue, respectively. Bars: (A–C, a) 10 µm; (A and B, b) 2 µm; (C, b and f) 200 nm.