XB-IMG-119660
Xenbase Image ID: 119660
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Figure 1. POM121-GFP and GFP-Nup153 label single NPCs and induce AL in live cells. (A–G) Images are confocal z-stacks (A and E) or single sections (B–D, F, and G). (A) Three-dimensional reconstruction of a live NRK cell nucleus expressing POM121-EGFP3. Top, maximum intensity projected of optical sections of the lower nuclear surface (double-pointed arrow). Inset enlarges 1 μm2 to show labeling of single NPCs. Bottom, xz slice at the line indicated in the top panel. (B) Live PtK2 cell coexpressing POM121-YFP3 (red) and SRβ-ECFP (green). AL labeled by POM121 are in direct contact to or colocalize with ER tubules and sheets as marked by SRβ. (C) HeLa cell expressing POM121-EGFP3 (green), fixed and stained for the endogenous nucleoporin p62 (red). Since the p62 antibody did not cross-react in PtK2 cells, HeLa cells were used for this experiment. AL containing POM121 colocalize with p62 antibodies (yellow in the merged lower panel, compare patterns in the split inset images). (D) Live PtK2 cell coexpressing ECFP–lamin B1 (green) and POM121-YFP3 (red). AL labeled by POM121 do not contain lamin B1. (E) Three-dimensional reconstruction of a live NRK cell nucleus expressing EGFP3-Nup153. Projections as in A. (F) Live COS7 cell coexpressing EGFP3-Nup153 (red) and SRβ-ECFP (green). COS7 cells were used for better ER morphology than NRK cells (compare with B). Yellow shows colocalization of AL with ER tubules. (G) COS7 cell expressing EGFP3-Nup153 (red) fixed and stained for the en-dogenous nucleoporin POM121 (red). AL containing Nup153 also label with anti-POM121 antibodies (yellow in the merged lower panel, compare patterns in the split inset images). Some POM121-positive structures do not contain Nup153. (H and I) Cryoimmuno double labeling electron micrographs from HeLa cells expressing POM121-EGFP3. Membrane boundaries are outlined next to the micrographs. Labeled are rat POM121 (10-nm gold) and human p62 (5-nm gold). POM121 and p62 colocalize on fenestrated membrane stacks (AL) in H and nuclear pores in I. M, mitochondria; N, nucleus; C, cytoplasm. Bars: (A–G) 5 μm; (H and I) 200 nm. Image published in: Daigle N et al. (2001) Copyright © 2001, The Rockefeller University Press. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike license Larger Image Printer Friendly View |