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. 2015 May 4;33(3):285–298. doi: 10.1016/j.devcel.2015.02.017

Figure 3.

Figure 3

Nup1 Overproduction Induces Nuclear Membrane Deformation

(A) Fluorescence imaging of cells expressing Nup188-GFP or Sec63-GFP (ER marker) and plasmid-borne Nup1 (1-123)-mCherry (GAL promoter). Induced refers to 4 hr of protein overproduction in galactose-containing medium. Black arrows indicate the site of nuclear membrane expansion; white arrows label NPC clusters. For protein expression levels, see Figure S3B. Scale bar represents 2 μm.

(B) Transmission electron microscopy (TEM) of representative cells, which overproduced Nup1 (1-123)-mCherry for 2, 4, or 8 hr, respectively. N, nucleus; C, cytoplasm; E, expansion membrane; V, vacuole. White arrows label single NPCs; black arrows mark the tip of the expansion membrane (2 hr) and newly formed intranuclear membrane sheets (8 hr). Scale bar represents 1 μm. Inset shows higher magnification of aligned NPCs; scale bar represents 100 nm.

(C) Live imaging of cells overexpressing the indicated Nup1 constructs for 4 hr in a NUP188-GFP background. See Figure S4B for Nup1 expression levels. Black arrows indicate expansion membrane; white arrows label NPC clusters. Note that Nup1 (1-76) overexpression in nup133Δ cells results in a similar NE phenotype, albeit with a more pronounced NPC clustering (Figure S3D). Scale bar represents 2 μm.

(D) TEM analysis of the indicated Nup1 fragments after 4 hr of overproduction. White arrows label single NPCs, same nomenclature as in (B). Scale bar represents 1 μm.