Extended Data Fig. 2: Defect in NPC and other non-core protein assembly persists in interphase MN in multiple cell lines.
From: Nuclear envelope assembly defects link mitotic errors to chromothripsis
a–c, Defective non-core NE protein recruitment to MN in RPE-1 cells. a, Top, experimental scheme. Bottom, representative images (quantified in b) with MN (arrows in enlarged images). Red letters, core NE proteins; green, non-core proteins. b, Quantification of results from a. The FI ratio of the indicated NE proteins in intact MN (RFP–NLS positive) relative to PN at the indicated time points after release from nocodazole block (mean with 95% CI, n = 97, 120, 106, 92, 114, 104, 113, 116, 104, 114, 121, 91, 114, 89, 113, 116, 125, left to right, from two experiments). Scale bars, 10 µm. c, MN/PN FI ratio of LAP2β in RPE-1 cells at the indicated time points after release from nocodazole block (mean with 95% CI, n = 70, 49, from two experiments). d, Deficiency of non-core proteins persists in MN in HeLa K (left) and U2OS cells (right). MN/PN FI ratio of the indicated NE proteins after release from nocodazole block as in a (mean with 95% CI, n = 79, 84, 111, 90, 79, 84, 111, 90, left to right for HeLa K, from two experiments; n = 53, 47, 45, 53, 66, 23, 64, 53, 27, 66, 54, 28, 53, 47, 46, 53, 69, 23, left to right for U2OS, from two experiments). e, Representative images of U2OS cells from d showing reduced assembly of B-type lamins in MN (arrows) 90 min after nocodazole release. Scale bars, 10 µm. f, Reduced NPC assembly on MN. MN/PN FI ratio of NPC proteins in RPE-1 cells 15 h after release from nocodazole block (mean with 95% CI, n = 47, 44, 49, 51, 49, 56, left to right, from two experiments). g, Reduced accumulation of B-type lamins but not A-type lamins on intact MN. Left, representative images of RPE-1 (left) and HeLa K (right) cells. In the enlarged merged images from the orange boxed region (HeLa K), the intensity of lamin B1 has been scaled differently to illustrate two points. First, there is reduction of lamin B but not lamin A/C in MN, which becomes evident when lamin B and lamin A/C on the main nucleus are scaled to the same intensity. Second, some MN display a lamin B1 gap, as has been reported6 (arrowheads indicate lamin B1 gap formed where the lamin A/C rim is continuous), which becomes evident when the lamin B1 intensity is scaled to a higher level. Right, general reduction of B-type lamins in MN. MN/PN FI ratio for the indicated lamins. Shown is the background-subtracted raw data for each individual micronucleus analysed in the indicated cell lines (n = 116 for RPE-1, n = 111 for HeLa K, from b, d above), 5 h after nocodazole release. Scale bars, 10 µm. A fraction of MN exhibit reduced lamin A/C, possibly owing to impaired import of lamin A/C during interphase.
