Spatial transcriptomics in the adult Drosophila brain and body
- VIB-KU Leuven Center for Brain and Disease Research, KU Leuven, Belgium
- Laboratory of Computational Biology, Department of Human Genetics, KU Leuven, Belgium
- Aix Marseille University, CNRS, IBDM, Turing Centre for Living Systems, France
- VIB Center for AI & Computational Biology, KU Leuven, Belgium
Figures
Figure 1 with 3 supplements
Principal workflow of spatial fly transcriptomics.
(A) Overview of the spatial transcriptomics workflow: adult flies were sectioned, sections were analyzed with Molecular Cartography and data were annotated using cell segmentation, rasterization …
Figure 1—figure supplement 1
Expression of the here selected marker genes in single-cell datasets.
(A) Heatmap showing expression in standardized log(CPM) of the 100 marker genes used in the head dataset in the single-cell clusters. (B) Density plot showing levels of gene expression detected in …
Figure 1—figure supplement 2
Head samples.
(A) All detected mRNA molecules color-coded in all 13 head samples used in the analysis. Scale bars represent 100 µm. Background stain labels DAPI. (B) Boxplot showing gene expression (number of …
Figure 1—figure supplement 3
Body samples.
(A) All detected mRNA molecules color-coded for all body samples used in the analysis. Sections are from the same male. Anterior is to the left, dorsal is up on each section and the section are …
Figure 2 with 3 supplements
Adult body cell types.
(A) Major cell types of adult males identified by marker genes. Scale bars represent 100 µm. Inset for gut shows zoom-ins of different regions (a: apical, b: basal). Inset for flight muscle, shows …
Figure 2—figure supplement 1
Motor neuron markers in flight muscles.
(A, B) Two different thorax sections are shown on the left and on the right. Molecular Cartography of neuronal markers in flight muscles (A). Molecular Cartography of neurotransmitter genes in …
Figure 2—figure supplement 2
Comparison of body spatial datasets with body single-cell datasets.
(A) Composite heatmap showing gene-gene co-expression based on Pearson correlation. Bottom triangle calculated on spatial datasets (using grid-based 5 µm squares). Top triangle calculated using …
Figure 2—figure supplement 3
Colocalization of gene expression in the body datasets.
(A) Overview of colocalization algorithm. For each mRNA spot a disk of 4 µm diameter was used as search space. Overlaps in the disk area were then used to calculate proximity between genes as …
Figure 3 with 1 supplement
Molecular Cartography shows mRNA-specific nuclear enrichment.
(A) Molecular Cartography (MC) visualization of marker genes of muscle subtypes. White boxes mark zoom-in regions shown in (B) and Figure 3—figure supplement 1. (B) Zoom-in on flight, leg, and head …
Figure 3—figure supplement 1
Subcellular mRNA localization in leg and head muscles.
(A) Molecular Cartography of sls, Act79B, Tpn41C and Mhc mRNAs in leg muscles, showing nuclei enrichment for sls mRNA and proximity of Act79B mRNA to leg muscle nuclei. (B) Molecular Cartography of s…
Figure 4 with 1 supplement
sls mRNA shows nuclear enrichment with increased concentration close to muscle-tendon junctions.
(A) Molecular Cartography visualization of sls mRNA (yellow) as pan muscle maker (same section as in Figure 3). White boxes mark zoom-in regions shown in (B) and (C). (B, C) Zoom-in on indirect …
Figure 4—figure supplement 1
Nuclear and sls mRNA localization.
(A) Molecular Cartography of sparsely expressed genes (salm and CG32121) at the anterior (right) and middle (left) flight muscle regions labeled in Figure 4A (see also the quantification of nuclear …
Figure 5 with 1 supplement
TpnC4, Act88F, and Mhc mRNAs flight muscle patterning.
(A) Molecular Cartography visualization of TpnC4, Act88F, and Mhc mRNAs on an adult fly section (same section as in Figure 3). The white box marks the zoom-in region shown in (B). (B) Zoom-in on the …
Figure 5—figure supplement 1
Variation in mRNA localization using Hybridization chain reaction-fluorescence in situ hybridization (HCR-FISH).
(A) HCR-FISH of TpnC4, Act88F, and sls mRNAs in an adult fly thorax. The white box marks the zoom-in region shown in (B). (B) Zoom-in of HCR-FISH of TpnC4, Act88F (blue in overlay), and sls mRNAs in …
Figure 6 with 1 supplement
Adult head cell types.
(A) tSNE showing expression of photoreceptor (ninaC), neuronal (para) and glial (repo) markers (left). Molecular Cartography of the same marker genes (right). (B, C) Molecular Cartography of marker …
Figure 6—figure supplement 1
Comparison of head spatial datasets with brain single-cell datasets.
(A) Gene-gene correlation measured across grid-based 5 µm squares and cells. Example mismatches shown in red, matching co-expression in green. (B) Composite heatmap showing gene-gene co-expression …
Figure 7 with 1 supplement
Comparison of different techniques for annotating the adult head samples.
(A) Overview of different spatial analysis methods which were used to annotate Molecular Cartography with labels from single-cell RNA-seq: grid-based, neighborhood embedding and nuclei segmentation. …
Figure 7—figure supplement 1
Tangram on grid-based quantification.
(A-D) Comparison of annotation of spatial data with single-cell RNA-seq. Grid-based squares are colored based on mapped single-cell cluster labels for (A) glia, (B) optic lobe, (C) central brain, …
Author response image 1
Barplots showing total number of mRNA molecules detected in Molecular Cartography (MC, Resolve, spatial spots) and in snRNA-seq data from the Fly Cell Atlas (10x Genomics, UMIs).
Individual black dots show individual experiments, counts are only shown for the chosen gene panel for each sample. Bar shows the mean, with error bars representing the standard error.
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| strain, strain background (Drosophila melanogaster) | Luminy | Loreau et al., 2023 | ||
| gene (Drosophila melanogaster) | sls | http://flybase.org/reports/FBgn0086906 | FBgn0086906 | |
| gene (Drosophila melanogaster) | Act88F | http://flybase.org/reports/FBgn0000047 | FBgn0000047 | |
| gene (Drosophila melanogaster) | Act88F | http://flybase.org/reports/FBgn0000047 | FBgn0000047 | |
| gene (Drosophila melanogaster) | TpnC4 | http://flybase.org/reports/FBgn0033027 | FBgn0033027 | |
| gene (Drosophila melanogaster) | Mhc | http://flybase.org/reports/FBgn0264695 | FBgn0264695 | |
| chemical compound, drug | OCT | VWR | 00411243 | |
| chemical compound, drug | Gelatin | Merck | porcine skin, 300 g Bloom Type A | |
| software, algorithm | SCANPY | Wolf et al., 2018; https://github.com/scverse/scanpy | RRID:SCR_018139 Version: v.1.8.1 | |
| software, algorithm | Tangram | Biancalani et al., 2021; https://github.com/broadinstitute/Tangram | Version: v1.0.2 | |
| software, algorithm | SpaGE | Abdelaal et al., 2020; https://github.com/tabdelaal/SpaGE | ||
| software, algorithm | Spage2Vec | Partel and Wählby, 2021; https://github.com/wahlby-lab/spage2vec | Version: v2.0 | |
| software, algorithm | Cellprofiler | Stirling et al., 2021; https://cellprofiler.org/ | RRID:SCR_007358 Version: v4.0.7 | |
| software, algorithm | SpatialNF | https://github.com/aertslab/SpatialNF |
Additional files
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MDAR checklist
- https://cdn.elifesciences.org/articles/92618/elife-92618-mdarchecklist1-v1.pdf
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Supplementary file 1
All 50 genes used for body MC experiments, including their expression levels and top expressing cell types inferred from the FCA.
- https://cdn.elifesciences.org/articles/92618/elife-92618-supp1-v1.xlsx
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Supplementary file 2
All 100 genes used for head MC experiments, including their expression levels and top expressing cell clusters inferred from the Pech et al., 2024.
- https://cdn.elifesciences.org/articles/92618/elife-92618-supp2-v1.xlsx
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Supplementary file 3
Detailed description of the 13 head section samples.
- https://cdn.elifesciences.org/articles/92618/elife-92618-supp3-v1.xlsx
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Supplementary file 4
All probe sequences used for the HCR-FISH experiments.
- https://cdn.elifesciences.org/articles/92618/elife-92618-supp4-v1.xlsx
