Single-cell transcriptomics of the Drosophila wing disc reveals instructive epithelium-to-myoblast interactions
- Department of Molecular and Cell Biology, University of California, Berkeley, United States
- Department of Electrical Engineering & Computer Science, Center for Computational Biology, UC Berkeley, University of California, Berkeley, United States
Figures
Figure 1 with 6 supplements
Temporal cell atlas of the developing wing-imaginal disc.
(A) Timeline of Drosophila development: embryo (E), larval phases (L1–L3), pupa, and adult. Diagram of the wing-imaginal disc within the third larval instar (L3) from mid (96 hr) and late (120 hr) …
Figure 1—figure supplement 1
Cell-type identification and temporal changes within major cell types.
(A) Dot plot summarizing gene expression for known markers of each of the identified cell types. For a gene of interest, the size of the dot indicates the percent of cells that express the gene and …
Figure 1—figure supplement 2
Temporal cell atlas of the wing disc epithelium.
(A) UMAP of epithelium cells, colored by cluster identities (same as in Figure 1H). Dot plot showing the level and percent of differentially-expressed (false discovery rate [FDR] < 0.05) marker gene …
Figure 1—figure supplement 3
Proximodistal axis stratifies the data more than anteroposterior axis.
(A–C) UMAPs of disc epithelium single-cell data showing expression levels of anterior compartment marker ci (A) and posterior compartment markers hh (B) and en (C). (D) Dot plot summarizing …
Figure 1—figure supplement 4
Correction of cell sex stratification within adult muscle precursor (AMP) scRNAseq data.
(A) UMAP of AMP single-cell data, as processed by our standard computational pipeline (without correction for cell sex or cell cycle). Colors correspond to 12 computational cell clusters as …
Figure 1—figure supplement 5
Correction of cell cycle stratification within adult muscle precursor (AMP) scRNAseq data.
(A–C) UMAPs of AMP single-cell data without any data correction (first column), after cell sex correction only (second column) (see Materials and methods and Figure 1—figure supplement 4 for …
Figure 1—figure supplement 6
Temporal changes within the direct and indirect adult muscle precursors (AMPs).
(A) Dot plot summarizing expression over time of genes that are differentially expressed within direct- or indirect-flight muscle precursors. Genes are grouped in the following manner: (1) genes …
Figure 2 with 1 supplement
Spatial mapping of single-cell data to a virtual wing disc and analysis of receptor–ligand expression.
(A) Schematic describing the creation of a three-layered virtual wing disc (adult muscle precursors [AMPs], disc proper [DP], and the peripodial epithelium [PE]) using DistMap (Karaiskos et al., 2017…
Figure 2—figure supplement 1
Mapping of epithelium cell clusters to the virtual wing disc.
(A) UMAP of epithelium cell clusters (same as the UMAP in Figure 1H). (B) Visualization of where epithelium cell clusters best map to the reference model for the disc proper layer (on left) and the …
Figure 3
Fibroblast growth factor (FGF) signaling between adult muscle precursors (AMPs) and epithelium is critical for AMP viability and numbers.
(A) FGF signaling pathway diagram. FGF ligands (Pyr and Ths) signal through the FGFR (Htl). (B–D') Expression of htl (B, B'), ths (C, C'), and pyr (D, D') in the single-cell data. (B), (C), and (D) …
Figure 4 with 1 supplement
Fibroblast growth factor (FGF) from disc epithelium controls adult muscle precursor (AMP) localization.
(A-B') Wing discs with dpp-Gal4 driving the expression of >GFP alone (A, A') or >GFP together with >pyr (B, B'), stained with anti-Twi (cyan) to visualize the AMPs. (A’) and (B’) correspond to white …
Figure 4—figure supplement 1
Ectopic Ths expression increases adult muscle precursor (AMP) number and migration.
(A, A') Wing discs with dpp-Gal4 driving the expression of both >GFP and >ths, stained with anti-Twi to visualize the AMPs. (A’) corresponds to the white dashed boxes in (A). Orthogonal section …
Figure 5 with 2 supplements
Hh signal from the disc epithelium patterns a subset of adult muscle precursors (AMPs).
(A–D) UMAPs of ptc (A), smo (B), ci (C), and hh (D) expression in AMPs. (E–E'') Notum of wing discs stained for anti-Ptc (red) with the hh-expressing epithelium cells marked by hh-Gal4 driving >GFP …
Figure 5—figure supplement 1
Ptc-expressing adult muscle precursors (AMPs) are neighboring the Hh-producing posterior compartment of the disc proper.
(A–A'''') Notum of wing discs stained for anti-Ptc (red) with 15B03-Gal4 driving >GFP. Magenta arrowheads in (A’’) indicate posterior-localized AMPs that display high Ptc expression. Orthogonal …
Figure 5—figure supplement 2
Adult wing-posture phenotypes and morphology of individual muscle fibers after Hh-signaling perturbation.
(A) Quantification of observed wing posture phenotypes under conditions of Hh-signaling perturbation within AMPs (for description of wing posture phenotypes, see Figure 5). All UAS lines are driven …
Figure 6 with 1 supplement
Nrt and Mid are novel downstream Hh-pathway targets in the adult muscle precursors (AMPs).
(A–C) UMAPs of Nrt (A), mid (B), and ptc (C) expression at 96 and 120 hr in AMPs. Note the increase in expression for Nrt and mid from 96 to 120 hr, whereas ptc expression is relatively unchanged. (D…
Figure 6—figure supplement 1
Knockdown of smo does not affect Ct protein levels in adult muscle precursors (AMPs).
(A–B') Notum of wing discs stained for anti-Cut (red) with 15B03-Gal4 driving >GFP alone (A, A') or >GFP with >smoRNAi (B, B'). Yellow arrowheads indicate higher levels of Ct staining in direct …
Author response image 1
FLAG-tagged Ths ligand is observed in both the epithelium notum and underlying AMPs.
(A-C) Max projections over all images slices (epithelium + AMPs) from wing discs with FLAG tag inserted at the N-terminus of Ths (Bl. 77476). Single image slice of the disc proper (DP) notum (D-F) …
Additional files
-
Supplementary file 1
Genes with differential expression between 96and 120 hr within the epithelium and adult muscle precursors (AMPs).
Genes were selected based on being significantly and consistently upregulated or downregulated between the two time points in either the disc epithelium and/or the AMPs. The average gene expression within cells (natural-log scale), fraction of cells expressing a given gene, fold-change between time points (natural-log scale), and false discovery rate (FDR) for differential expression significance are reported. These gene expression, detection, and fold-change calculations are averaged across each of the pairwise comparisons performed, and the max FDR value is shown (see Materials and methods for details on differential expression between time points). Negative fold-change values indicate higher expression at 96 hr and are colored magenta. Positive fold-change values indicate higher expression at 120 hr and are colored green. N.R.: not replicable; calculations in which the fold-change direction differed between pairwise comparisons.
- https://cdn.elifesciences.org/articles/61276/elife-61276-supp1-v2.xlsx
-
Supplementary file 2
Genes with differential expression between 96 and 120 hr within the epithelial cell clusters.
Genes were selected based on being significantly and consistently upregulated or downregulated between the two time points in at least one epithelial cluster. The natural-log of the fold-change between 96 and 120 hr is reported, averaged across each of the pairwise comparisons performed (see Materials and methods for details on differential expression between time points). Negative values indicate higher expression at 96 hr and are colored magenta. Positive values indicate higher expression at 120 hr and are colored green. Values that were not significant (based on max false discovery rate) are reported with a ‘-'.
- https://cdn.elifesciences.org/articles/61276/elife-61276-supp2-v2.xlsx
-
Supplementary file 3
Genes with differential expression between 96 and 120 hr within the direct and indirect adult muscle precursors (AMPs).
Genes were selected based on being significantly and consistently upregulated or downregulated between the two time points in either the direct and/or the indirect AMPs. The average gene expression within cells (natural-log scale), fraction of cells expressing a given gene, fold-change between time points (natural-log scale), and false discovery rate (FDR) for differential expression significance are reported. These gene expression, detection, and fold-change calculations are averaged across each of the pairwise comparisons performed, and the max FDR value is shown (see Materials and methods for details on differential expression between time points). Negative fold-change values indicate higher expression at 96 hr and are colored magenta. Positive fold-change values indicate higher expression at 120 hr and are colored green. N.R.: not replicable; calculations in which the fold-change direction differed between pairwise comparisons.
- https://cdn.elifesciences.org/articles/61276/elife-61276-supp3-v2.xlsx
-
Supplementary file 4
Geometry of disc model.
CSV file of the X, Y, Z geometry used in reference gene expression patterns (Supplementary file 5). Formatted as used in DistMap to generate virtual wing disc.
- https://cdn.elifesciences.org/articles/61276/elife-61276-supp4-v2.csv
-
Supplementary file 5
Reference gene expression patterns.
CSV file of the binarized reference gene expression patterns (along with geometry in Supplementary file 4). Formatted as used in DistMap to generate virtual wing disc.
- https://cdn.elifesciences.org/articles/61276/elife-61276-supp5-v2.csv
-
Transparent reporting form
- https://cdn.elifesciences.org/articles/61276/elife-61276-transrepform-v2.pdf
