Med19^AID removal results in the deregulation a limited part of the wing imaginal disc transcriptome.

(A) Scheme depicting the experimental setup used to produce wing imaginal discs from ub>TIR1 Med19AID larvae in which Med19AID degradation has been triggered, or not (no Auxin control- see GFP signal), upstream of the RNA-seq analysis. (B) Differential gene expression analysis volcano plot showing the expression fold change after Med19AID degradation and associated adjusted p-values. 8753 genes found expressed in the wing imaginal disc are depicted. Genes with significative fold change (adjusted p-value 2FC 2FC >0,5).</p

The expression of most of canonical housekeeping (HK) genes is not affected by Med19AID depletion.

Scatter plot analysis of the expression fold change (Log2FC) of genes belonging to several families of housekeeping functions (x axis, list detailed in S3 Table). The red dashed lines mark Log2FC = +/ - 0.5. (TIF)</p

Med19 promotes the expression of the spatially regulated developmental genes.

(A) Scatter plot depicting the distribution of the Log2FC values of the 573 transcription factors found significantly expressed in the wing imaginal disc from the RNAseq data. The transcription factors with a demonstrated spatial transcriptional regulation (SR TFs) in the wing imaginal disc are shown in purple (n = 122). (B) Dot plot 10x10 showing the proportion of the non-regulated (dark grey), upregulated (red), or downregulated (blue) genes among the 122 SR transcription factors. (C) Bar graph made from RT-qPCR data showing the relative level of transcripts of 6 genes known to be spatially regulated (wg, cut, E(Spl)m8, sal or ubiquitous (Myosin, E2F1) in the wing discs of ub>; Med19AID-GFP larvae after Med19AID depletion was carried out for 4 hours in all the disc cells (percentage of the control in which Med19AID degradation was not triggered). The graph shows the mean and standard deviation of 3 independent experiments. (D) Confocal images of the wing pouch region of dissected en>; Med19AID wing discs that were subjected to Med19AID degradation (en> 3H Aux), or not (control: No Gal4 driver, and en> no Aux), and in which wingless (wg) pre-mRNA were visualized by smiFISH using intronic probes and Med19 degradation by direct GFP fluorescence. top images:Med19AID GFP signal. bottom images: wg smiFISH signal. (E) Wing disc D/V boundary posterior to anterior wg smiFISH signal ratio established in en>; Med19AID wing discs: No auxin control (no Aux. n = 5), 1H auxin (n = 6), and 3H auxin (n = 6). ****: P value <0.0001 (Unpaired t test). (F) Hierarchical clustering tree of the 30 best FDR of the GO terms associated with the downregulated genes. The size of the solid circles is proportional to the enrichment False Discovery Rate (FDR). GO clusters appear related to development (purple), nucleic acid synthesis (blue), and cell cycle regulation (yellow).</p

List of the 58 Drosophila tissues and developmental stages (modENCODE project) used to establish the RNA expression profile of the genes expressed in the wing imaginal disc.

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30 best GO terms of the down-regulated genes sorted by FDR enrichment obtained on ShinyGO web site.

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List of canonical housekeeping genes.

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Model, derived from the analysis of the nature of the genes deregulated or not after Med19^AID degradation, showing how Med19 may function.

(A) Med19 acts as a critical cofactor of transcription factors whose activity is regulated by signaling pathways, in negative or positive control of transcription. (B) Med19 is not a Mediator subunit required for the expression of the constitutively expressed genes.</p

Venn diagram showing the overlap between HK—HK* (top), and SR—SR*(bottom) lists of genes.

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The expression of <i>achaete</i> (<i>ac</i>) and <i>wingless</i> (<i>wg</i>) is impaired specifically in the domain of the wing imaginal disc where Med19AID is degraded.

(A) Confocal images of ap>TIR1; Med19AID wing imaginal disc immunostained for Ac and GFP (Med19AID detection). Upper row: Control wing disc in which Med19AID degradation did not occur (no driver). Bottom row: Wing disc in which Med19 degradation was induced for 3 hours in the dorsal compartment, using an apterous driver. Right column: Magnification in the anterior wing pouch Ac staining pattern emphasizing the loss of Ac expression in the dorsal row of cells of the anterior wing margin when Med19AID degradation is triggered in the dorsal compartment of the wing disc. (B) en>TIR1; Med19AID wing imaginal discs immunostained for Wg and GFP (Med19AID), in which the degradation of Med19AID was operated with the en> driver for 3 hours (bottom row), or not triggered (control, upper row). (TIF)</p

Original uncropped and unadjusted images for blots or gels from Figs 1A and 5D.

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