Journal: Communications biology

Article Title: Depletion of SUN1/2 induces heterochromatin accrual in mesenchymal stem cells during adipogenesis.

doi: 10.1038/s42003-025-07832-3

Figure Lengend Snippet: Fig. 1 | SUN1/2 depletion alters nuclear mor- phology. a Representative images of MSCs treated with siRNA targeting SUN1/2 (siSUN) which were stained for SUN1 (green), SUN2 (red), and DNA (blue). b siSUN-treated cells had 47% decrease of SUN1 intensity (n = 1478, P < 0.0001). c SUN2 intensity levels were decreased by 52% in siSUN- treated cells (n = 1478, P < 0.0001). d MSCs treated with siSUN had an increase in nuclear area by 7% (n = 1478, p 0 < 0.01). e Nucleus perimeter decreased in siSUN-treated MSCs by 8% (n = 1478, p < 0.0001). f Nuclear circularity decreased by 9% in siSUN-treated cells (n = 1478, p < 0.001). Compar- isons were made against control using non- parametric Mann–Whitney test where *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. The scale bar represents 50 μm. Results were presented as mean ± STD.

Article Snippet: The plasmids used for the knockdown-rescue experiments are: pEGFP-SUN1_785 (Addgene 125852), SUN2 (NM_001199579) Human Tagged ORF Clone (Origene RC234801), and pCMV6-Entry (OriGene PS100001) as the empty vector control. qPCR About 2 ul of each CUT&RUN sample was run in a 20 ul reaction following Bio-Rad protocols targeting Adipoq (Bio-Rad, 10025636).

Techniques: Staining, Control, MANN-WHITNEY

Journal: Scientific reports

Article Title: The SUN2-nesprin-2 LINC complex and KIF20A function in the Golgi dispersal.

doi: 10.1038/s41598-021-84750-4

Figure Lengend Snippet: Figure 6. Working model for the regulation of the Golgi organization by LINC complex. Left, The minus-end- directed MT motor activity, i.e., dynein (blue), dominates plus-end-directed motor activity under a steady-state condition, and juxtanuclear accumulation of the Golgi complex is observed. Right, In SUN1-depleted cells, KIF20A is activated or suppression of KIF20A activity is cancelled, and KIF20A activity dominates dynein activity. An explanation for the inhibition of KIF20A activity in the presence of SUN1 is that LINC complex formations are biased towards SUN1 inclusion and SUN2/nesprin-2 LINC complex formation is suppressed. In response to SUN1 depletion, incorporation of nesprin-2 in the SUN1-LINC complex decreased and SUN2/ nesprin-2 LINC complexes are formed. As a result, functions of KIF20A acting with a SUN2/nesprin-2 LINC complex becomes predominant (orange) over the minus-end-directed motor activities (blue), and the Golgi complex is dispersed.

Article Snippet: Myc-tagged mouse SUN1 encoding 913 amino acids (NM_024451) and Myc-tagged mouse SUN2 were purchased from Origene (Rockville, MD, USA).

Techniques: Activity Assay, Inhibition

Journal: Molecular Biology of the Cell

Article Title: Fluorescence fluctuation spectroscopy reveals differential SUN protein oligomerization in living cells

doi: 10.1091/mbc.E17-04-0233

Figure Lengend Snippet: SUN2 oligomerization in the NE. (A) Constructs used in this figure. (B) Represent­ative epifluorescence images of U2OS cells expressing the indicated constructs. Scale bar: 5 μm. (C–E) Plots of b vs. N for the indicated constructs. The data in C were fitted to a monomer/dimer/trimer binding model (solid red line), which is shown in D and E (dashed red line), with K MD = 4100 (1000 μM) , K DT = 0.06 (0.01 μM) , and a monomer/trimer binding model (solid green line) with K MT = 26 (6 μM) ± 7.

Article Snippet: NC (TR30015), SUN1 (TF300647B/FI302582), and SUN2 (TF300646A/FI302577) HuSH-29 shRNA constructs in pRFP-C-RS were purchased from OriGene Technologies).

Techniques: Construct, Expressing, Binding Assay

Journal: Molecular Biology of the Cell

Article Title: Fluorescence fluctuation spectroscopy reveals differential SUN protein oligomerization in living cells

doi: 10.1091/mbc.E17-04-0233

Figure Lengend Snippet: SUN1 and SUN2 oligomerization in the cytoplasm. (A) Constructs used in this figure. (B) Representative epifluorescence images of U2OS cells expressing the indicated constructs. Scale bar: 5 μm. (C–G) Plots of b vs. N for the indicated constructs. The data in C were fitted to a trimeric binding model (solid blue line), which is shown in D and E (dashed blue line) with K MD = 8000 (60 μM) ± 4000 and K DT = 0.3 (0.002 μM) ± 0.2. The data in F were fitted to a monomer/trimer/hexamer binding model (solid blue line) with K MT = 100 (0.7 μM) ± 60 and a trimer–hexamer dissociation coefficient K TH = 1500 (10 μM) ± 400, which is then shown in G (dashed blue line). Estimated binding curves (dashed red lines) for the data obtained in the NE for the indicated constructs are presented in C and F by converting N from the NE to its cytoplasmic value.

Article Snippet: NC (TR30015), SUN1 (TF300647B/FI302582), and SUN2 (TF300646A/FI302577) HuSH-29 shRNA constructs in pRFP-C-RS were purchased from OriGene Technologies).

Techniques: Construct, Expressing, Binding Assay

Journal: Molecular Biology of the Cell

Article Title: Fluorescence fluctuation spectroscopy reveals differential SUN protein oligomerization in living cells

doi: 10.1091/mbc.E17-04-0233

Figure Lengend Snippet: Models of SUN1 and SUN2 oligomerization. (A) Working model of SUN2 oligomerization. (B’ and B’’) Working models of SUN1 oligomerization with the NE, which lead to the assembly of higher-order SUN1 oligomers (SUN1 N ).

Article Snippet: NC (TR30015), SUN1 (TF300647B/FI302582), and SUN2 (TF300646A/FI302577) HuSH-29 shRNA constructs in pRFP-C-RS were purchased from OriGene Technologies).

Techniques:

Journal: Molecular Biology of the Cell

Article Title: Fluorescence fluctuation spectroscopy reveals differential SUN protein oligomerization in living cells

doi: 10.1091/mbc.E17-04-0233

Figure Lengend Snippet: Primers used to generate the constructs used in this paper.

Article Snippet: NC (TR30015), SUN1 (TF300647B/FI302582), and SUN2 (TF300646A/FI302577) HuSH-29 shRNA constructs in pRFP-C-RS were purchased from OriGene Technologies).

Techniques: Construct, Sequencing

Journal: JCI Insight

Article Title: Multiomic analysis of microRNA-mediated regulation reveals a proliferative axis involving miR-10b in fibrolamellar carcinoma

doi: 10.1172/jci.insight.154743

Figure Lengend Snippet: ( A ) Protein expression of DNAJB1-PRKACA (DP) is detected with a protein kinase A catalytic α subunit (PKA) antibody. WT PKAc, DP major, and DP minor are identified. Lane 1, FLC-H cell line; lane 2, nonmalignant liver; lane 3, FLC patient sample; lane 4, FLC PDX sample. Vinculin expression for loading control is shown in the lower panel. Uncropped immunoblot shown in , A and B. ( B ) qPCR showing the RQV of miR-10b in FLC-H cells 6 days after 500 nM treatment with miR-10b LNA or scrambled sequence compared with mock ( n = 4 each condition). ( C ) Luciferase signal (RLU) in FLC-H cells after 6 days of 500 nM miR-10b LNA treatment is shown as RQV compared with the scrambled negative control (6 trials, n = 6 each condition). ( D ) qPCR showing the RQV of FANCC , KLF11 , SEC14L2 , SIRT5 , SUN2 , and TRIM35 in FLC-H cells after 6 days of 500 nM miR-10b LNA treatment compared with the negative control ( n = 5–7 trials with 3 replicates for each condition, SIRT5 n = 2 trials). ( E ) Soft agar colony formation of FLC-H cells 35 days after 500 nM miR-10b LNA compared with the negative control shown as RQV. ( F ) Representative nitro blue tetrazolium–stained images shown (2 trials, n = 8 each condition). ( G ) EdU incorporation in FLC-H cells 6 days after 500 nM treatment with miR-10b LNA compared with the negative control shown as RQV (2 trials, n = 6 each condition). ( H ) Representative DAPI- and EdU-stained images show total and proliferative cells, respectively. Scale bars: 100 μm. In all assays, each dot represents the average signal across technical replicates for a single biological replicate. P values are calculated by 2-tailed Student’s t test. P values reported in B and D were adjusted for multiple testing correction post hoc by the Benjamini-Hochberg method.

Article Snippet: Individual gene assay IDs include the following: CDH1, hs01023895; DNAJB1-PRKACA, custom; FANCC, hs0098454; KLF11, hs00231614; miR-10b, 002218; miR-21, 000397; miR-182, 002334; PTEN, hs02621230; RNU6, 001973; RPS9, hs02339424; SEC14L2, hs00391446; SIRT5, hs00978331; SUN2, hs00391446; and TRIM35, hs00324633 (Thermo Fisher Scientific).

Techniques: Expressing, Control, Western Blot, Sequencing, Luciferase, Negative Control, Staining

Journal: The Journal of Cell Biology

Article Title: Nuclear envelope rupture is induced by actin-based nucleus confinement

doi: 10.1083/jcb.201603053

Figure Lengend Snippet: The LINC complex is absent from chromatin hernia. Representative images of U2OS GFP-NLS shLmnB1 cells arrested with thymidine, fixed, and labeled with LmnA antibodies (top three rows, rabbit; bottom, mouse) and (from top to bottom) Sun1, Sun2, nesprin1 (Nsp1), and nesprin2 (Nsp2) antibodies. Bars, 10 µm.

Article Snippet: Antibodies used for immunoblotting were as follows: mouse antitubulin (1:5,000; Sigma-Aldrich), mouse anti-GAPDH (1:5,000; Genetex), rabbit anti-Sun1 (1:1,000; Novus Biologicals), and rabbit anti-Sun2 (1:1,000; Genetex).

Techniques: Labeling