Journal: Biochemical and biophysical research communications

Article Title: Ascl1-mediated enhancement of GABAergic neuronal function in differentiated F11 cells under high glucose conditions.

doi: 10.1016/j.bbrc.2025.151721

Figure Lengend Snippet: Fig. 2. Ascl1 overexpression enhances GABAergic neuronal function in differentiated F11 cells. (A) Timeline for lenti-

Article Snippet: Lentivirus vector generation The full-length open reading frame of Ascl1 was obtained from plasmid #27150 (Addgene), corresponding to Ascl1 (NM_008553).

Techniques: Over Expression

Journal: Biochemical and biophysical research communications

Article Title: Ascl1-mediated enhancement of GABAergic neuronal function in differentiated F11 cells under high glucose conditions.

doi: 10.1016/j.bbrc.2025.151721

Figure Lengend Snippet: Fig. 3. Regulation of GABAergic gene expression and inflammatory responses by Ascl1 in a high-glucose (HG)

Article Snippet: Lentivirus vector generation The full-length open reading frame of Ascl1 was obtained from plasmid #27150 (Addgene), corresponding to Ascl1 (NM_008553).

Techniques: Gene Expression

Journal: Nature communications

Article Title: Cenpj/CPAP regulates progenitor divisions and neuronal migration in the cerebral cortex downstream of Ascl1.

doi: 10.1038/ncomms7474

Figure Lengend Snippet: Figure 1 | Ascl1 directly regulates Cenpjexpression in the embryonic telencephalon. (a) Illustration of the Ascl1-binding signal in E14.5 telencephalic cells obtained by chromatin immunoprecipitation with promoter microarrays (chromatin immunoprecipitation (ChIP)-chip). The plot displays the ChIP enrichment ratio for Ascl1 (red) and control (black) samples in promoter regions. Black arrow indicates the transcription start site and direction of transcription. (b,c) Western blot analysis of Cenpj protein levels extracted from E14.5 Ascl1 mutants compared with wild-type (WT) telencephalon. Data presented as mean±s.e.m., n ¼ 3, Student’s t-test, *Po0.05. (d,e) Immunohistochemistry for Cenpj on coronal sections of WT and Ascl1 mutant E14.5 embryos. The Cenpj protein (d) is localized with the centrosome marker centrin in apical cortical progenitors and the signal is reduced in Ascl1 mutant progenitors (e). Scale bars, 1 mm. (f) Quantitative PCR analysis of Cenpj transcripts extracted from E14.5 Ascl1 mutants compared with WT telencephalon. Data presented as mean±s.e.m., n ¼ 3, Student’s t-test, *Po0.05. (g–i) In situ hybridization for Cenpj on coronal sections of the developing telencephalon at E14.5 from WT (g,i) and Ascl1 knockout mouse (h) using an antisense Cenpj probe (g,h) and a control Cenpj sense probe (i). Expression of Cenpj was reduced in the ventricular zone and cortical plate of the Ascl1 mutant cortex. Scale bar, 100 mm. DAPI, 40,6-diamidino-2-phenylindole.

Article Snippet: Cenpj shRNA (50-TCCGACTGGATGCCTG GAAGAGAGCAGAA-30), Ascl1 shRNA (50-AGAAGATGAGCAAGGTGGAGA CGCTGCGC-30) control shRNA (50-GCACTACCAGAGCTAACTCAGATAGT ACT-30) plasmids were purchased from Origene.

Techniques: Binding Assay, Chromatin Immunoprecipitation, ChIP-chip, Control, Western Blot, Immunohistochemistry, Mutagenesis, Marker, Real-time Polymerase Chain Reaction, In Situ Hybridization, Knock-Out, Expressing

Journal: Nature communications

Article Title: Cenpj/CPAP regulates progenitor divisions and neuronal migration in the cerebral cortex downstream of Ascl1.

doi: 10.1038/ncomms7474

Figure Lengend Snippet: Figure 6 | Cenpj is the main effector of Ascl1for centrosome biogenesis. (a,b) Labelling of centrosomes in apical cortical progenitors from wild-type (WT) (a) or Ascl1 null embryos (b) with an antibody to g-tubulin. Insets to the right show higher magnification of the boxed areas with separate colour channels. Scale bars, 1 mm. (c) Quantification of the area of the centrosomes in WT and Ascl1 null embryos labelled with an antibody to g-tubulin. Three embryos analysed for each condition. Student’s t-test ***Po0.001. (d,e) Analysis of the centrosome area of GFP þ -dividing apical progenitors electroporated 1 day earlier in utero with Ascl1 shRNA (d) or an Ascl1 shRNA plus a pCMV-Cenpj construct (e) labelled for GFP (green), pH3 (red) and g-tubulin (grey). Insets to the right show higher magnification of the boxed areas with g-tubulin (grey) channels. Scale bars, 1 mm. (f) Quantification of the area of the centrosomes in control shRNA-electroporated progenitors, Cenpj shRNA, Ascl1 shRNA and Ascl1 shRNA þ pCMV-Cenpj-electroporated progenitors, labelled with g-tubulin. The centrosomes are smaller in Ascl1-silenced progenitors and this defect is rescued in Ascl1-silenced cells co-electroporated with a Cenpj expression vector. Student’s t-test ***Po0.001. (g,h) Percentage of cells electroporated with Ascl1 shRNA (g) and Ascl1 shRNA þ pCMV-Cenpj (h) having a vertical (60–90) or oblique (30–60) cleavage angle. Ascl1 shRNA, n ¼ 33 cells; Ascl1 shRNA þCenpj, n ¼ 24 cells. Cells were analysed from at least six embryos obtained from three litters. DAPI, 40,6-diamidino-2-phenylindole; NS, not significant.

Article Snippet: Cenpj shRNA (50-TCCGACTGGATGCCTG GAAGAGAGCAGAA-30), Ascl1 shRNA (50-AGAAGATGAGCAAGGTGGAGA CGCTGCGC-30) control shRNA (50-GCACTACCAGAGCTAACTCAGATAGT ACT-30) plasmids were purchased from Origene.

Techniques: In Utero, shRNA, Construct, Control, Expressing, Plasmid Preparation

Journal: Nature communications

Article Title: Cenpj/CPAP regulates progenitor divisions and neuronal migration in the cerebral cortex downstream of Ascl1.

doi: 10.1038/ncomms7474

Figure Lengend Snippet: Figure 7 | Cenpj is the main effector of Ascl1for microtubule regulation. (a–c) Analysis of radial migration 3 days after electroporation of E14.5 cortices with GFP and a control shRNA, Ascl1 shRNA or Ascl1 shRNA þ pCMV-Cenpj. Scale bar, 50 mm. (d) Quantification of the migration of GFP þ cells in the different zones of the cortex. The accumulation of Ascl1-silenced cells in the IZ but not their depletion from the uCP was rescued by overexpression of Cenpj. Student’s t-test *Po0.05 (e–g). Abnormal leading-process enlargement of Ascl1-silenced neurons in the CP. Scale bar, 5 mm. (h) Quantification of leading process thickness. Three embryos analysed for each condition; control shRNA, n ¼ 177 cells; Ascl1 shRNA, n ¼ 128 cells; Ascl1 shRNA þ pCMV-Cenpj, n ¼ 239 cells. Student’s t-test **Po0.01. (i–k) Nocodazole treatment of cultured neurons 3 days after electroporation. Ascl1 silencing preserved a higher level of acetylated tubulin labelling in the cell body and leading process than control shRNA, and overexpression of Cenpj restored the acetylated tubulin-labelling intensity. Scale bar, 5 mm. (l) Quantification of acetylated tubulin labelling co-localized with the nucleus. Three independent cultures analysed for each condition; control shRNA, n ¼ 40 cells; Ascl1 shRNA, n ¼ 53; Ascl1 shRNA þ pCMV-Cenpj, n ¼ 37. Student’s t-test *Po0.05; **Po0.01. DAPI, 40,6-diamidino-2-phenylindole; NS, not significant.

Article Snippet: Cenpj shRNA (50-TCCGACTGGATGCCTG GAAGAGAGCAGAA-30), Ascl1 shRNA (50-AGAAGATGAGCAAGGTGGAGA CGCTGCGC-30) control shRNA (50-GCACTACCAGAGCTAACTCAGATAGT ACT-30) plasmids were purchased from Origene.

Techniques: Migration, Electroporation, Control, shRNA, Over Expression, Cell Culture