plasmids  (Addgene inc)

Journal: Stem Cell Reports

Article Title: ASCL1 promotes nuclear shrinkage in transdifferentiation by suppressing NUP37

doi: 10.1016/j.stemcr.2026.102823

Figure Lengend Snippet: Reduction of nuclear size during the direct conversion of human fibroblasts to neurons (A) Schematic for the transdifferentiation of human fibroblasts to iNs by lentiviruses expressing ASCL1, miR124-9-9 ∗ -BclxL, and p53 shRNA (AMp, uppercase for overexpression and lowercase for knockdown). –FBS, serum withdrawal to synchronize cell cycle at the G1/S checkpoint. Scale bar, 100 μm. (B) Phase contrast images of MRC5 cells under conversion at the indicated time points. Scale bar, 100 μm. Insets, super-resolution images of DAPI-stained nuclei. Scale bar, 10 μm. (C) Nuclear volume quantification for MRC5 cells throughout reprogramming. n = 50 frames from 3 independent experiments for each time point, unpaired t test vs. day −2. ∗ p < 0.01. (D) Quantification of nuclear area for MRC5 cells throughout reprogramming. n = 50 frames from 3 independent experiments for each time point, unpaired t test vs. day −2, ∗ p < 0.01. (E) Quantification of nuclear area at the indicated time points as MRC5, AG22056 newborn foreskin fibroblasts, or GM09918 (78 years) skin fibroblasts were being converted to iNs. n = 50 frames from 3 independent experiments for each time point, unpaired t test vs. day −2, ∗ p < 0.01. (F) The average area of MRC5, AG22056, or GM09918 cells as fibroblasts at day −2 (Fib) or TUJ1 + or MAP2 + iNs. ns, no significance. n = 50 frames from three independent experiments. (G) iPSC-derived cortical neurons were co-stained for MAP2 and DAPI at days 30, 40, and 80 of differentiation. Scale bar, 50 μm. Inset, super-resolution images of neuronal nuclei; scale bar, 10 μm. (H) Quantification of nuclear area of the indicated samples. ∗ p < 0.01, vs. the preceding bar (or D30 for iPSC-derived neurons), n = 50 frames from 3 independent experiments.

Article Snippet: We purchased the following plasmids from Addgene: pLKO.1/ p53 shRNA (#19119), pLKO.1/scrambled shRNA (#1864), pMD2.G (#12259), psPAX2 (#12260), pTight-9-124-Bclx (miR9/9 ∗ -124, #60857), pRL-SV40P (#27163), and pGL3 enhancer vector (#212938).

Techniques: Expressing, shRNA, Over Expression, Knockdown, Staining, Derivative Assay

Journal: Stem Cell Reports

Article Title: ASCL1 promotes nuclear shrinkage in transdifferentiation by suppressing NUP37

doi: 10.1016/j.stemcr.2026.102823

Figure Lengend Snippet: NUP37 knockdown significantly enhanced AMp-mediated transdifferentiation and nuclear shrinkage (A) Western blot of NUP37 in MRC5 cells transduced without (−) or with the indicated reprogramming factors. (B–D) MRC5 human fibroblasts reprogrammed with ASCL1, MIR124-9-9 ∗ -BclxL, and p53 shRNA (AMp) (B), AMp and NUP37 shRNA (AMpu) (C), or AMp and NUP37 overexpression (AMpU) (D) were co-stained as indicated on day 14. Scale bar, 100 μm. (E–G) Reprogramming efficiency (E) as measured by the percentages of TUJ1 + or MAP2 + cells among all DAPI + cells, reprogramming yield of MAP2 + cells per frame (F), and the number of DAPI + cells per frame (G) at day 14. # and ∗ , p < 0.05, n = 15 (3 experiments, 5 frames each), vs. AMp for the indicated cell type, unpaired t test. (H) Nuclear area of MAP2 + neurons for each condition. ∗ p < 0.001, n = 50 frames from 3 independent experiments, vs. AMp, unpaired t test. (I–P) MRC5 cells reprogrammed with AMp (I–L) or AMpu (M–P) were co-stained as indicated at different time points. Scale bar, 100 μm. (Q‒S) (Q) Reprogramming efficiency of MAP2 + -generated neurons per DAPI + nuclei. (R) Yield of MAP2 + neurons. (S) Number of DAPI + cells per frame. ∗ p < 0.01, n = 15 (3 experiments, 5 frames each), vs. AMp at the same time point, unpaired t test. (T) RT-qPCR measurement of mature neuronal markers in AMp- or AMpu-induced neurons at D14. ∗ p < 0.05, n = 6 (3 experiments, duplicate for each), vs. AMp, unpaired t test.

Article Snippet: We purchased the following plasmids from Addgene: pLKO.1/ p53 shRNA (#19119), pLKO.1/scrambled shRNA (#1864), pMD2.G (#12259), psPAX2 (#12260), pTight-9-124-Bclx (miR9/9 ∗ -124, #60857), pRL-SV40P (#27163), and pGL3 enhancer vector (#212938).

Techniques: Knockdown, Western Blot, shRNA, Over Expression, Staining, Generated, Quantitative RT-PCR

Journal: Stem Cell Reports

Article Title: ASCL1 promotes nuclear shrinkage in transdifferentiation by suppressing NUP37

doi: 10.1016/j.stemcr.2026.102823

Figure Lengend Snippet: Cooperation of ASCL1 and NUP37 shRNA in reprogramming and nuclear shrinkage (A–P) MRC5 human fibroblasts were reprogrammed without or with the indicated combinations of ASCL1 (A), miR124-9-9 ∗ -BclxL (M), p53 shRNA (p) and NUP37 shRNA (u), and co-stained as indicated at day 14. Bar, 100 μm. (Q–S) Reprogramming efficiency (Q) as measured by the percentages of TUJ1 + or MAP2 + cells among all DAPI + cells, reprogramming yield of MAP2 + cells per frame (R), and the number of DAPI + cells per frame (S) at day 14. # and ∗ , p < 0.001, n = 15 (3 experiments, 5 frames each), vs. the corresponding condition without u for the indicated cell type, unpaired t test. $ p < 0.001, n = 15 (3 experiments, 5 frames each), vs. no virus (−V), unpaired t test. (T) Nuclear area for each condition. ∗ p < 0.05, n = 50 frames from 3 independent experiments, vs. the corresponding condition without u; unpaired t test. $ p < 0.005, n = 50 frames from 3 independent experiments, vs. no virus (−V), unpaired t test.

Article Snippet: We purchased the following plasmids from Addgene: pLKO.1/ p53 shRNA (#19119), pLKO.1/scrambled shRNA (#1864), pMD2.G (#12259), psPAX2 (#12260), pTight-9-124-Bclx (miR9/9 ∗ -124, #60857), pRL-SV40P (#27163), and pGL3 enhancer vector (#212938).

Techniques: shRNA, Staining, Virus