1. Volume 26, Issue 19, Pages 2651-2658 (October 2016)
A-type Lamins Form Distinct Filamentous Networks with Differential Nuclear Pore Complex Associations 
Wei Xie, Alexandre Chojnowski, Thomas Boudier, John S.Y. Lim, Sohail Ahmed, Zheng Ser, Colin Stewart, Brian Burke 
Current Biology 
Volume 26, Issue 19, Pages (October 2016)
DOI: /j.cub
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2. Current Biology 2016 26, 2651-2658DOI: (10.1016/j.cub.2016.07.049)
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3. Figure 1
BioID Analysis of LaA and LaC Reveals Differential Associations with the Nuclear Pore Complex Components Tpr and Nup214
(A) Confocal immunofluorescence microscopy of cells expressing BirA-LaA and BirA-LaC in the absence or presence of doxycycline (DOX) induction, showing the proper nuclear lamina localization of the BirA-LaA and BirA-LaC constructs.
(B) Western blot analysis of the same cells using antibodies against myc-BirA and GAPDH.
(C) Graphic representation of the interactome of selected proteins identified from the BioID experiment, and of their associations as identified by the STRING database ( The edge color of the nodes is mapped to the reliability of each identified protein (see the Supplemental Experimental Procedures), from high confidence (dark color) to low confidence (light color). The bar graphs on each node represent the relative quantity of its protein identified in each BioID experiment: BirA-LaA no dox (black), BirA-LaC no dox (white), BirA-LaA +dox (green), and BirA-LaC +dox (blue).
(D) Quantification of protein intensity (AU) identified by the BioID screen for the NPC proteins Tpr, Nup214, Nup155, Nup98, and Pom121 and proteins localizing to the nuclear lamina A-type lamins (Lmna), Emerin (Emd), Sun1, and Sun2. Values were mean ± SEM (n ≥ 3), ∗p < 0.05, ∗∗p < 0.01, two-way ANOVA with Tukey’s post hoc-test.
See also Table S1.
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4. Figure 2
Exogenously Expressed LaA and LaC Form Filamentous Networks at the Nuclear Envelope and Exhibit Distinct Associations with NPCs
(A and B) Lmna−/− MAFs expressing exogenous mEos2-tagged LaA (A) or LaC (B) proteins, respectively, were fixed and subjected to PALM super-resolution imaging with the microscope focused at the surface of the nucleus. Areas highlighted in the top panels were enlarged at the bottom, showing strings of lamin localizations.
(C–E) Lmna−/− MAFs expressing exogenous mEos2-tagged LaA (C), LaA-mature (D), and LaC (E), respectively, were fixed and labeled with the NPC-specific antibody QE5 and subjected to sequential dSTORM (green) and PALM (red) super-resolution imaging with the microscope focused at the nuclear surface.
(F) NPC association with respective A-type lamin variants were quantified, as percentage of NPCs localized inside the lamin occupied regions to the total number of NPCs within the individual cell nuclei. Values were mean ± SEM (n ≥ 10), statistical significance were analyzed by two-tailed Student’s t tests. ∗p < 0.05; ∗∗p < 0.01; ns not significant.
See also Figure S1.
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5. Figure 3
Endogenous Lamins Are Organized into Distinct Networks at the Nuclear Periphery
Lmna+/+ or Lmna−/− MAFs were fixed and co-labeled with antibodies against LaA, LaC, LaB1, and NPC as indicated.
(A and B) Top: individual channels from a super-resolved dSTORM image were analyzed and rendered using the EMST plugin in Fiji, with the computed network (upper right) super-imposed on endogenous localizations (upper left) of LaA and LaC (A), LaA and LaB1 (B). Bottom: areas highlighted in the top panels were enlarged, showing individual lamin channels as well as the corresponding merged image.
(C) dSTORM localizations of LaA (left, red), LaC (middle, red), or LaB1 (right, red) in Lmna+/+ MAFs, together with NPCs (green), were super-imposed with computed network of respective lamins (magenta).
(D) dSTORM localizations of LaB1 (red) and NPCs (green) in Lmna−/− MAFs were super-imposed with computed network of LaB1 (magenta).
See also Figure S2.
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6. Figure 4
Different Lamin Species Self-Associate to Form Independent and Exclusive Protein Complexes
(A and B) Left: Lmna+/+ (A) or Lmna−/− (B) MAFs were fixed and labeled with antibody against endogenous LaA (red) and LaB1 (green), followed by dSTORM super-resolution imaging focused at the surface of the nucleus; right: area highlighted in the left panel was enlarged, showing LaB1 localizations.
(C–E) N-terminally epitope tagged LaA, LaC, and LaB1 proteins were synthesized separately in vitro and then mixed together in pairs as indicated. Following co-immunoprecipitation using an antibody against the V5 tag, western blot analysis was performed employing antibodies against either V5, HA, or Myc tags. Arrow head indicates V5 antibody heavy chain.
See also Figure S3.
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