Journal: Nature Communications

Article Title: Conserved transcriptional regulation by BRN1 and BRN2 in neocortical progenitors drives mammalian neural specification and neocortical expansion

doi: 10.1038/s41467-024-52443-x

Figure Lengend Snippet: a Schematic of progenitor’s neurogenic mode in control (black) and Brn1/2-cKO (red). b UMAP of scRNAseq Tbr2 , MKi67 and Neurod2 expression. c UMAP of cells going through indirect and direct neurogenesis in control (CT) and Brn1/2-cKO (cKO) at E12.5 and E14.5. d Proportion of cells going through indirect and direct neurogenesis by age and genotype (Pearson’s Chi-squared test: E12.5-p = 2.212e−11, E14.5-p < 2.2e−16). e TBR2 (red) and Ki67 (grey) immunolabeling in control and Brn1/2-cKO cortical sections at E12.5 and E14.5 (E12.5: n = 5 CT, n = 4 cKO mice; E14.5: n = 3 mice/group; two-sided unpaired t- test: E12.5-p = 0.05, E14.5-p = 0.006). Boxed area at higher magnification on the right. f Volcano plot: differentially expressed genes (DEG) between Brn1/2-cKO progenitors and controls at E14.5 highlighting NOTCH signaling-associated genes (Monocle3 VGAM test; SD = 0.15; q < 0.05; Supplementary Data ). g ChIP-qPCR analysis of BRN1/2 binding to the indicated promoters/enhancers at E14.5 ( n = 3/condition; two-sided unpaired t-test: Notch1 -p = 0.0004, Dll1 -p = 0.0066, Hes1 -p = 0.0042). h HES1 (red) immunolabeling in the ventricular zone (VZ) of control and Brn1/2-cKO cortical sections at E14.5 ( n = 10 CT, n = 8 cKO mice; two-sided unpaired t- test: p < 0.0001). i In utero electroporation (IUE) in Brn1 fl/fl ; Brn2 fl/fl mice at E14.5. Cell identities of the control, Brn1/2-cKO and Brn1/2-cKO + NOTCH1 condition immunolabeled for RFP (red), TBR2 ( j , grey) and NEUROD2 ( k , grey) at E16.5. Boxed area: higher magnification in inserts. RFP + TBR2 + and RFP + NEUROD2 + at E16.5 in the control, Brn1/2-cKO , Brn1/2-cKO + NOTCH1 ( l ) and Brn1/2-cKO + DLL1 ( m ) condition (NOTCH1( l ): n = 8 CT, n = 8 cKO, n = 7 cKO+NOTCH1; DLL1( m ): n = 5 CT, n = 5 cKO, n = 6 cKO+NOTCH1; one-way ANOVA-Dunnett’s multiple comparisons test: NOTCH1-TBR2-p < 0.0001, F 2,20 = 20.50; NOTCH1-NEUROD2-p = 0.001, F 2,21 = 9.702; DLL1-TBR2-p = 0.0048, F 2,13 = 8.294; DLL1-NEUROD2-p = 0.0098, F 2,13 = 6.748). Low and top lines represent the limits of the VZ and cortical plate (CP), respectively. Lines and dashed lines outline cells expressing or lacking expression of the indicated marker, respectively. SVZ Subventricular Zone, IP Intermediate Progenitors, N Neurons. Values are mean ± SEM; ns, not significant; * p < 0.05, ** p < 0.01, *** p < 0.001. Scale bars: 50 µm (lower magnification), 10 µm (higher magnification). Source data are provided as a Source Data file.

Article Snippet: pCAG-mRFP (plasmid #32600, Addgene) was deposited by Anna-Katerina Hadjantonakis . pCAG-CRE (plasmid #13775, Addgene) was generated by Connie Cepko . pCS2 NOTCH1 Full Length-6MT (pCS2- Notch1-FL , plasmid #41728, Addgene) was deposited by Raphael Kopan . pCAG- Brn -DBD- EnR (plasmid #19714, Addgene) and pCAG- Brn2 (plasmid #19711, Addgene) were generated by Connie Cepko . pCMV6 Delta1 Full Length Myc-DDK was obtained from Origene (pCMV6- Dll1-FL , #MR226161). pBlue-hASPM(WT) was generated by Byoung-Il Bae.

Techniques: Control, Expressing, Immunolabeling, ChIP-qPCR, Binding Assay, In Utero, Electroporation, Marker

Journal: Nature Communications

Article Title: Conserved transcriptional regulation by BRN1 and BRN2 in neocortical progenitors drives mammalian neural specification and neocortical expansion

doi: 10.1038/s41467-024-52443-x

Figure Lengend Snippet: a In utero electroporation (IUE) in wild-type ferrets at E35. Cell identities of the control (CT) and Brn-Enr ( B-Enr ) condition immunolabeled for RFP (red), Ki67 ( b , grey), TBR2 ( c , grey) and NEUROD2 ( d , grey) at E37 ( n = 4 CT, n = 7 Brn-Enr ferrets; two-sided unpaired t- test: Ki67-p = 0.0014, TBR2-p = 0.0012, NEUROD2-p = 0.0004). f NOTCH1 (grey ) expression in RFP + cells of the electroporated ventricular zone (VZ) ( n = 4 ferrets/group; two-sided unpaired t- test: p = 0.0099). g ASPM or CDK6 (grey) expression in RFP + cells of the electroporated cortex ( n = 3 CT, n = 4 Brn-Enr ferrets; two-tailed unpaired t- test: ASPM -p = 0.0006, CDK6 -p = 0.0001). h RFP + cells expressing OLIG2 (gray) in the electroporated cortex ( n = 4 CT, n = 5 Brn-Enr ferrets; two-sided unpaired t- test: p = 0.0009). h’ Total OLIG2 + cells in the electroporated cortex ( n = 4 CT, n = 5 Brn-Enr ferrets; two-sided unpaired t- test: p = 0.0226). i ScRNAseq DATA from E36 cortex of control (CT) and BRN2 KO (KO) cynomolgus monkey re-analyzed using the same pipeline we used for the analysis in mice. UMAP of scRNAseq from control and BRN2 KO by genotype ( j ) and cell type ( k ). l UMAP of gene signatures for apical progenitors (AP; e.g., NES ), basal progenitors (BP; e.g., BTG2 ) and neurons (e.g., DCX ) in control and BRN2 KO. m UMAP of BP signature (1), MKi67 expression (2) and TRIcycle score (3) in control and BRN2 KO. n UMAP of cells going through indirect and direct neurogenesis in control and BRN2 KO. o Proportion of cells going through indirect and direct neurogenesis by genotype (Pearson’s Chi-squared test: p = 4.454e-12). p Expression of NOTCH signaling-associated genes in control and BRN2 KO progenitors. q Expression of the indicated cortical upper layer (UL) and deep layer (DL) neuronal marker genes in control and BRN2 KO neurons. r Expression of primary microcephaly DEG in BRN2 KO progenitors compared to controls. Boxed areas: higher magnification in inserts. Lines and dashed lines outline cells expressing or not expressing the indicated marker, respectively. iSVZ Inner Subventricular Zone, oSVZ Outer Subventricular Zone. Values are mean ± SEM; * p < 0.05, ** p < 0.01, *** p < 0.001. Scale bars: 50 µm (lower magnification), 10 µm (higher magnification). Source data are provided as a Source Data file.

Article Snippet: pCAG-mRFP (plasmid #32600, Addgene) was deposited by Anna-Katerina Hadjantonakis . pCAG-CRE (plasmid #13775, Addgene) was generated by Connie Cepko . pCS2 NOTCH1 Full Length-6MT (pCS2- Notch1-FL , plasmid #41728, Addgene) was deposited by Raphael Kopan . pCAG- Brn -DBD- EnR (plasmid #19714, Addgene) and pCAG- Brn2 (plasmid #19711, Addgene) were generated by Connie Cepko . pCMV6 Delta1 Full Length Myc-DDK was obtained from Origene (pCMV6- Dll1-FL , #MR226161). pBlue-hASPM(WT) was generated by Byoung-Il Bae.

Techniques: In Utero, Electroporation, Control, Immunolabeling, Expressing, Two Tailed Test, Marker

Journal: Cell reports

Article Title: Notch receptor-ligand binding facilitates extracellular vesicle-mediated neuron-to-neuron communication

doi: 10.1016/j.celrep.2024.113680

Figure Lengend Snippet:

Article Snippet: Notchl-myc , Addgene/Raphael Kopan lab, Schroeter et al. , Cat# 41728; RRID:Addgene_41728.

Techniques: Virus, Recombinant, Protease Inhibitor, Lactate Dehydrogenase Assay, Magnetic Beads, Silver Staining, Peptide Fractionation, RNA Sequencing Assay, Mass Spectrometry, Software, Sequencing, Transmission Assay, Imaging, Microscopy