Journal: bioRxiv

Article Title: Translatome analysis reveals cellular network in DLK-dependent hippocampal glutamatergic neuron degeneration

doi: 10.1101/2024.07.10.602846

Figure Lengend Snippet: (A) Western blot validation of immunoprecipitation of HA-tagged Rpl22. (B) qRT-PCR expression of Vglut1 (glutamatergic neuron marker), Wfs1 (CA1 marker), Vgat (inhibitory neurons), Gfap (astrocytes) shows expression of transcripts expressed in glutamatergic neurons and depletion of non-glutamatergic neuron transcripts. N=3 biological replicates, data shown as fold change of expression of marker genes in immunoprecipitated glutamatergic neuron RNA relative to whole hippocampal RNA. Normalized to Gapdh expression. (C) Venn diagram showing overlap of genes with statistically significant differential translation in DLK(cKO) and DLK(OE). (D,E) Heatmaps of significant genes from DLK(cKO) and DLK(OE). Columns represent individual mice expression levels; rows represent individual genes with the right-hand labels showing which dataset the gene was found to be statistically significant in. Data were normalized by row, with color keys shown above the heatmap. (F,G,H,I) Pie charts show expression of differentially expressed genes based on adult, endogenous expression patterns in the Allen Mouse Brain Atlas in situ data. (F,H) Upregulated, or (G,I) downregulated genes when DLK expression is (F,G) increased or (H,I) conditionally knocked out are categorized based on expression patterns in CA1, CA3, and DG in dorsal hippocampus. (J) Sunburst plot shows significant enrichment for differentially expressed genes from DLK(cKO) relating to the synapse. (K) Pathway analysis of expression data for mice with increased DLK compared to control. Nodes represent sets of genes involved in pathways, with size dependent on the number of genes in the pathway. Nodes are clustered in shaded circles based on related pathways. All pathways shown have q-value (false discovery rate <0.05). Red represents pathways enriched in mice with increased DLK. Blue indicates pathways downregulated in mice with increased DLK.

Article Snippet: Gene expression patterns of differentially translated genes were evaluated using Allen Mouse Brain Atlas in situ data from P56 mice, and supplemented with data from through the Single cell portal from the Broad Institute or data from , adolescent data through mousebrain.org when no in situ data was available or when expression was weak.

Techniques: Western Blot, Biomarker Discovery, Immunoprecipitation, Quantitative RT-PCR, Expressing, Marker, In Situ, Control

Journal: bioRxiv

Article Title: Translatome analysis reveals cellular network in DLK-dependent hippocampal glutamatergic neuron degeneration

doi: 10.1101/2024.07.10.602846

Figure Lengend Snippet: (A) Volcano plot showing RiboTag analysis of gene expression in Vglut1 Cre/+; H11-DLK iOE/+ ;Rpl22 HA/+ vs Vglut1 Cre/+ ;Rpl22 HA/+ (age P15). 260 genes showing differential expression with adjusted p-values < 0.05 are shown in red. Names of genes with p<1E-10 are labeled. (B) Volcano plot showing RiboTag analysis of genes in Vglut1 Cre/+ ;DLK(cKO) fl/fl ;Rpl22 HA/+ vs Vglut1 Cre/+ ;Rpl22 HA/+ (age P15). 36 genes showing differential expression with adjusted p-values < 0.05 are shown in blue. Names of genes with p<1E-10 are labeled. (C) Rank-rank hypergeometric overlap comparison of gene expression in DLK(iOE) and DLK(cKO) datasets shows enrichment of similar genes when DLK is low or high. Color represents the -log transformed hypergeometric p-values (blue=weaker p-value, red=stronger p-value). (D,E) Gene ontology (GO) analysis of significantly upregulated or downregulated genes, respectively, when DLK expression is increased in hippocampal glutamatergic neurons. Colors correspond to P-values. Circle size represents fold enrichment for the GO term, with number on X position showing # of significant genes included in the GO term. (F) SynGO sunburst plot shows significant enrichment for differentially expressed genes when DLK expression is increased in hippocampal glutamatergic neurons. (G,H) Pie charts show distribution of synaptic genes whose expression exhibits dependency when DLK expression is increased in hippocampus, based on in situ data in CA1, CA3, and DG in dorsal hippocampus (P56) in the Allen Mouse Brain Atlas.

Article Snippet: Gene expression patterns of differentially translated genes were evaluated using Allen Mouse Brain Atlas in situ data from P56 mice, and supplemented with data from through the Single cell portal from the Broad Institute or data from , adolescent data through mousebrain.org when no in situ data was available or when expression was weak.

Techniques: Gene Expression, Quantitative Proteomics, Labeling, Comparison, Transformation Assay, Expressing, In Situ

Journal: Cerebral Cortex (New York, NY)

Article Title: A Layer-specific Corticofugal Input to the Mouse Superior Colliculus

doi: 10.1093/cercor/bhx161

Figure Lengend Snippet: Areal characterization of cortico-superior-collicular neurons. (a) Epifluorescence images show retrogradely labeled neurons in a series of slices containing the AC. Dashed yellow lines indicate areal boundaries between the dorsal, primary, and ventral AC. (b) Schematic from the Allen Institute for Brain Science coronal mouse atlas which was used to reference the boundaries between the cortical areas. (c) Continuing in series from panel a. (d) Continuing in series from panel b.

Article Snippet: Dashed yellow lines indicate areal boundaries between the dorsal, primary, and ventral AC. ( b ) Schematic from the Allen Institute for Brain Science coronal mouse atlas which was used to reference the boundaries between the cortical areas. ( c ) Continuing in series from panel a. ( d ) Continuing in series from panel b.

Techniques: Labeling

Journal: Communications Biology

Article Title: Targeted volumetric single-molecule localization microscopy of defined presynaptic structures in brain sections

doi: 10.1038/s42003-021-01939-z

Figure Lengend Snippet: a View of a virtual mouse brain with the dimensions of the hippocampal formation in green and a horizontal section from Allen Mouse Brain Atlas (image credit: Allen Institute for Brain Science, available from: http://mouse.brain-map.org/static/brainexplorer ) to demonstrate the hippocampal region used for imaging (white box). b Confocal image of mossy fibers in a 25 µm thick hippocampal slice from a Thy1mEGFP(Lsi1) mouse. The white box marks again the region of interest (25 µm × 25 µm × 25 µm) that is shown magnified in the next panel. c 3D volume reconstruction of mEGFP in xyz -view, showing axons and mossy fiber boutons (green), a typical bouton with filopodial extensions is marked and further highlighted in d . The window (10 µm in z ) for en bloc imaging in the 25 µm thick tissue block is shown in blue. d 3D volume reconstruction of a mossy fiber bouton. The light green area of the bouton shows the volume that would be captured using a typical 1 µm imaging depth in z for a single focal plane. The white box marks a virtual synaptic contact that is further magnified in the next panel. e A virtual synaptic contact in side view (presynaptic membrane in green, Bassoon clusters in magenta, Homer-1 in yellow, postsynaptic membrane in gray), scale bar in b 200 µm, in e 100 nm.

Article Snippet: Fig. 2 Two-color imaging of hippocampal mossy fibers and Bassoon clusters along the ventro-dorsal axis. a Ventral view of a virtual mouse brain with landmarks for trimming in yellow (left) and horizontal sections from Allen Mouse Brain Atlas (image credit: Allen Institute for Brain Science, available from: https://mouse.brain-map.org/static/brainexplorer ) to illustrate the ventral (level 1, middle panel) and the 600 μm more dorsal region (level 2). b Confocal image of mossy fibers in a Thy1EGFP(M) mouse with zinc-transporter 3 (magenta) staining.

Techniques: Imaging, Blocking Assay, Membrane

Journal: Communications Biology

Article Title: Targeted volumetric single-molecule localization microscopy of defined presynaptic structures in brain sections

doi: 10.1038/s42003-021-01939-z

Figure Lengend Snippet: a Ventral view of a virtual mouse brain with landmarks for trimming in yellow (left) and horizontal sections from Allen Mouse Brain Atlas (image credit: Allen Institute for Brain Science, available from: https://mouse.brain-map.org/static/brainexplorer ) to illustrate the ventral (level 1, middle panel) and the 600 µm more dorsal region (level 2). b Confocal image of mossy fibers in a Thy1EGFP(M) mouse with zinc-transporter 3 (magenta) staining. A white box marks the imaging window (30 µm × 30 µm) shown enlarged in the next panel. c Two-color d STORM image of synapses stained for presynaptic Bassoon (green) and postsynaptic Homer 1 (magenta). White box indicates magnified view in the next panel. d , e Synaptic contacts in different orientations: upper box—side view, lower box—plane view further magnified in e . f Protein domain layout of Mus musculus Bassoon and Homer1: Zinc-finger (Zn), coiled coil (CC), PxxP, and enabled/VASP homology (EVH); epitopes of the anti-Bassoon (green) and anti-Homer 1 (magenta) antibodies. Histograms of length ( g ) and counts ( i ) of Bassoon clusters in side view at level 1 (blue, n = 181 clusters in nine images from eight animals) and level 2 (magenta, n = 208 clusters in eight images from eight animals). Clusters at level 2 are longer ( p < 0.001) and have more counts ( p < 0.05) than at level 1. Summary box plots (horizontal line median, boxes 25th and 75th, whiskers 10th and 90th percentile) in all eight individual animals for length ( h ) and localization counts ( j ) (* p < 0.05; ** p < 0.01; *** p < 0.001). i Scale bars 100 µm ( b ), 3 µm ( c ), 1 µm ( d ), and 200 nm ( e ).

Article Snippet: Fig. 2 Two-color imaging of hippocampal mossy fibers and Bassoon clusters along the ventro-dorsal axis. a Ventral view of a virtual mouse brain with landmarks for trimming in yellow (left) and horizontal sections from Allen Mouse Brain Atlas (image credit: Allen Institute for Brain Science, available from: https://mouse.brain-map.org/static/brainexplorer ) to illustrate the ventral (level 1, middle panel) and the 600 μm more dorsal region (level 2). b Confocal image of mossy fibers in a Thy1EGFP(M) mouse with zinc-transporter 3 (magenta) staining.

Techniques: Staining, Imaging