Single-cell transcriptomics identifies Keap1-Nrf2 regulated collective invasion in a Drosophila tumor model
- Department of Biochemistry and Molecular Biology, Tulane University School of Medicine, Tulane Cancer Center, United States
- Department of Biological Science, Florida State University, United States
Figures
Figure 1 with 3 supplements
Inducing Lgl knockdown in follicle cells causes distinct phenotypic and transcriptomic changes.
(A) Left: orthogonal projection of a single ovariole displaying individual egg chambers containing experimental control follicle cells at early and midoogenesis. Follicle cells at mitotic stages are …
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Figure 1—source data 1
Raw data for (unnormalized) IntDen values of RFP, GFP, and DAPI enrichment.
Related to Figure 1C.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig1-data1-v2.xlsx
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Figure 1—source data 2
Quantification of the stage-specific phenotypic characterization of ovarioles containing egg chambers with 96 hr Lgl-KD in their follicle cells.
Related to Figure 1D.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig1-data2-v2.xlsx
Figure 1—figure supplement 1
Lgl loss of function in follicle cells causes invasive multilayering and cell fate heterogeneity.
(A) Confocal images of tjTS-driven Lgl-KD ovaries (right) compared to that of a tjTS experimental control (left). Inducing Lgl-KD in follicle cells causes late-stage egg chambers to abort germline …
Figure 1—figure supplement 2
Whole-tissue RNA-seq of samples containing multilayered Lgl-KD follicle cells.
(A) Representative confocal images of representative ovarioles from the four samples used as input for RNA-seq analysis. Nucleus is marked by DAPI (white). Scale bars: 50 µm. (B) Enriched Gene …
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Figure 1—figure supplement 2—source data 1
Differentially expressed genes in 96h-Lgl-KD vs. others comparison.
Related to Figure 1—figure supplement 2B.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig1-figsupp2-data1-v2.xlsx
Figure 1—figure supplement 3
Single-cell RNA-seq of ovaries with 72h-Lgl-KD follicle cells.
(A) Scatter plots depicting the correlation between the different RNA-seq samples. Red triangle in the left plot represents markers from late development stages of oogenesis, representing the …
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Figure 1—figure supplement 3—source data 1
Cluster-specific markers of the tjTS>lglRNAi (72 hr) single-cell RNA-seq (scRNA-seq) dataset.
Related to Figure 1—figure supplement 3B.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig1-figsupp3-data1-v2.xlsx
Figure 2
Integration of w1118 and tjTS>lglRNAi single-cell datasets identifies sample-specific clusters.
(A) UMAP plot of 17,874 w1118 follicle cells integrated with 12,923 tjTS>lglRNAi follicle cells, grouped into 20 clusters, with their approximated identities listed below. (B) Canonical marker …
Figure 3 with 2 supplements
Unique clusters of tjTS>lglRNAi dataset exhibit distinct gene expression and regulon activity.
(A) Left: UMAP plot of the re-embedded tjTS>lglRNAi follicle cells with the unique clusters being highlighted. Right: RNA velocity vectors superimposed on the embedded cells of unique tjTS>lglRNAi …
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Figure 3—source data 1
Regulon specificity scores (RSS) of regulons enriched in clusters 7, 8, and 13 in tjTS>lglRNAi (72 hr) single-cell RNA-seq (scRNA-seq) dataset.
Related to Figure 3F.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig3-data1-v2.xlsx
Figure 3—figure supplement 1
Validation of overlapping markers in Lgl-KD cells.
(A) Gene enrichment plot for Xbp1 expression (red) is shown on top left. Expression of UPR stress sensor Xbp1-GFP is observed in clusters representing early stages of development as well as clusters …
Figure 3—figure supplement 2
Expression of JNK signaling components in 72h-Lgl-KD cells.
(A) Confocal images showing antibody staining against pJNK (green) in multilayered egg chambers containing RFP+ (red) Lgl-KD follicle cells. Nucleus is marked by DAPI (white). F-actin is marked by …
Figure 4 with 1 supplement
Cluster 7 cells exhibit heterogenous gene expression and regulon activity.
(A) UMAP plot of subdivided cluster 7 cells, superimposed with RNA velocity vectors. (B) Heatmap representing regulon–regulon correlation based on Connection Specificity Index (CSI) of active …
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Figure 4—source data 1
Regulon specificity scores (RSS) of regulons enriched in the subclusters of cluster 7 in tjTS>lglRNAi (72 hr) single-cell RNA-seq (scRNA-seq) dataset.
Related to Figure 4C.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig4-data1-v2.xlsx
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Figure 4—source data 2
Differentially expressed markers of the subclusters of cluster 7 cells in tjTS>lglRNAi (72 hr) single-cell RNA-seq (scRNA-seq) dataset.
Related to Figure 4D.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig4-data2-v2.xlsx
Figure 4—figure supplement 1
GstD-lacZ expression in Lgl-KD+Keap1-OE follicle cells and w1118 experimental control.
(A) Confocal image of a multilayered egg chamber containing Lgl-KD+Keap1-OE follicle cells (green). GstD-lacZ expression, marked by β-gal staining (red), is observed spuriously in the multilayered …
Figure 5 with 1 supplement
Keap1-Nrf2 signaling drives invasiveness of Lgl-KD multilayers.
(A) Representative confocal images of ovarioles containing egg chambers with transgene expression in follicle cells (green) driving Lgl-KD (top), Lgl-KD+Cnc-KD (middle), and Lgl-KD+Keap1-KD (bottom …
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Figure 5—source data 1
Quantification of the rescue of stage-specific Lgl-KD phenotypes upon Cnc-KD and Keap1-KD.
Related to Figure 5B.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig5-data1-v2.xlsx
Figure 5—figure supplement 1
Epithelial multilayering and invasiveness are increased in Lgl-KD+CncC-OE egg chambers.
(A) Confocal images showing increased invasiveness of the epithelia containing Lgl-KD+CncC-OE follicle cells (green). Nurse cell (yellow arrow) is depicted being compressed by invading …
Figure 6 with 1 supplement
Invading Lgl-KD+Keap1-OE cells exhibit extensive cytoskeletal remodeling.
(A) Confocal images of egg chambers with Lgl-KD (left) and Lgl-KD+Keap1-OE (middle and right panels) in the follicle cells (red; shown in the middle row). Egg chambers are stained with Shg (DE-Cad; …
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Figure 6—source data 1
Related to Figure 6B.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig6-data1-v2.xlsx
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Figure 6—source data 2
Raw data for the IntDen values of F-actin enrichment in the monolayered and multilayered follicle cells expressing Lgl-KD and Lgl-KD+Keap1-OE.
Related to Figure 6D.
- https://cdn.elifesciences.org/articles/80956/elife-80956-fig6-data2-v2.xlsx
Figure 6—figure supplement 1
Oxidative stress response is not detected in cluster 7 cells.
(A) Confocal images showing dihydroethidium (DHE) staining (red) in the multilayers. Significant DHE staining is not observed within the multilayered cells. Nucleus is marked by DAPI (white). …
Additional files
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MDAR checklist
- https://cdn.elifesciences.org/articles/80956/elife-80956-mdarchecklist1-v2.docx
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Source data 1
Enriched transcription factor binding motifs detected in genes expressed in the cells of cluster 7.
- https://cdn.elifesciences.org/articles/80956/elife-80956-data1-v2.xlsx
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Source data 2
Genes showing dynamic behavior of transcription driving the underlying RNA velocity of cluster 7 cells.
- https://cdn.elifesciences.org/articles/80956/elife-80956-data2-v2.xlsx
