EMBO/soda-nel huggingface dataset:
Tissue resident macrophages actively suppress Il-1β release via a reactive prostanoid/IL-10 pathway (2020)
Figure 5

Fig.5 The aberrant IL-1β production by Gata6-KOmye pMɸ is not led by mitochondria dysfunction.

(A) Flow cytometry analysis of Mitotracker (MT) Green and Red integration in Gata6-WT and Gata6-KOmye naïve pMɸ (gated on F4/80hi/+ Tim4+) in vivo, 30 min after intraperitoneal injection (i.p.) of 1 µM of Mitotracker Green and Red. Data shown are representative of 3 to 5 independent mice of each genotype and are expressed as mean +/- SEM. Student's t test analysis was performed.

(B) Flow cytometry analysis of Mitotracker green and red staining in Gata6-WT and Gata6-KOmye pMɸ (gated on F4/80hi/+ Tim4+) unstimulated (-) or after LPS (100 ng.ml-1), IL-10 (10 ng.ml-1), αIL-10R (5 µg.ml-1) or isotype (5 µg.ml-1) stimulation for 16 h in vitro. Data shown are pooled from 3 independent experiments and normalized to WT unstimulated.

(C) Flow cytometry analysis of MitoSOX staining in Gata6-WT and Gata6-KOmye pMɸ (gated on F4/80hi/+ Tim4+) unstimulated (-) or after LPS (100 ng.ml-1), IL-10 (10 ng.ml-1) and αIL-10R (5 µg.ml-1) stimulation for 16 h in vitro. Data shown are pooled from 2 independent experiments and normalized to WT unstimulated.

(D) IL-1β ELISA of supernatants of Gata6-WT and Gata6-KOmye pMɸ unstimulated (-) or stimulated with 100 ng.ml-1 LPS, 500 µM MitoTempo, 10 mM NAC, 0.5 µM MitoQ for 16 h. Data are representative of at least 3 experiments.

Data information: Data are expressed as mean +/- SEM and two-way ANOVA statistical analysis with Tukey's multiple comparison post-test were performed unless otherwise stated. *p<0.05, ***p<0.001
Tissue resident macrophages actively suppress Il-1β release via a reactive prostanoid/IL-10 pathway (2020)
Figure 6

Fig.6 PGI2-derived IL-10 controls IL-1β processing.

(A) mRNA expression analysis of Il1b, Nlrp3 and Caspase1 (Casp1) of Gata6-WT and Gata6-KOmye pMФ stimulated for 3 h with 100 ng.ml-1 LPS, 10 µM beraprost, 10 ng.ml-1 IL-10, 5 µg.ml-1 αIL-10R or 5 µg.ml-1 isotype. Data are representative of at least 3 independent experiments.

(B) Western blot analysis (left) and quantification (right) of Gata6-WT and Gata6-KOmye pMФ stimulated for 6 h with 100 ng.ml-1 LPS, 10 µM beraprost, 10 ng.ml-1 IL-10 or 5 µg.ml-1 αIL-10R. Data are representative of 3 independent experiments.

(C) Caspase1 activity analysis of Gata6-WT and Gata6-KOmye pMФ stimulated for 16 h with 100 ng.ml-1 LPS, 10 µM beraprost, 10 ng.ml-1 IL-10, 5 µg.ml-1 αIL-10R or 5 µg.ml-1 isotype. n = 5 to 8 individual mice.

(D) Mean fluorescence intensity (MFI) of pro-IL-1β of Gata6-WT and Gata6-KOmye pMФ stimulated for 3 h with 100 ng.ml-1 LPS, 10 µM beraprost, 10 ng.ml-1 IL-10, 5 µg.ml-1 αIL-10R or 5 µg.ml-1 isotype for 16 h or freshly isolated. n = 6 to 13 individual mice. Data were log transformed before performing statistical analysis.

(E) mRNA expression analysis of Il10 and Tnf of Gata6-WT and Gata6-KOmye pMФ stimulated for 3 h with 100 ng.ml-1 LPS, 10 µM beraprost, 10 ng.ml-1 IL-10 or 5 µg.ml-1 αIL-10R. Data are representative of at least 3 independent experiments.

(F) Representation of the mechanism of action of the Gata6-PGI2-IL-10 active inhibitory signal on IL-1β processing.

Data information: Data are expressed as mean +/- SEM and two-way ANOVA statistical analysis with Tukey's multiple comparison post-test was performed. *p<0.05, **p<0.01, ***p<0.001
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 1

Figure 1. Bax translocation to mitochondria correlates with mitochondrial depolarization.

(A) Confocal live cell imaging of HeLa cells overexpressing GFP-Bax (green) and mitochondria marked with TMRE 100 nM (magenta) show cytosolic GFP-Bax distribution and healthy mitochondria. Lower panels are zoomed images corresponding to the white rectangle in the upper panel and represent the individual and merged channels. Time shows the minutes after apoptosis induction with Staurosporine (STS) 1 µM. Scale bars, 5µm (overview) and 2µm (zooms)

HeLa cells shown in A at longer times after STS treatment. Images show sequential GFP-Bax translocation (green), which correlates with mitochondria depolarization (TMRE, magenta). Scale bars, 5 µm

HeLa cells overexpressing GFP-Bax mutant (1-2/L-6) (green) and mitochondria marked with TMRE 100 nM (magenta). Images show constitutive GFP-Bax localization to healthy mitochondria without apoptosis induction. Right panels are zoomed images corresponding to the white rectangle in the left panel and represent the individual and overlaid channels. Scale bars, 5µm (overview) and 2 µm (zooms)

Standard deviation of the fluorescence intensity of GFP-Bax blue) versus the corrected total cell fluorescence (CTCF) of TMRE (purple) in individual cells (N=4). CTCF= Integrated Density – (Area of selected cell X Mean fluorescence of 3 background readings). Time 0 corresponds to the normalized time when both events cross in each cell. Black line represents the average of the individual cells.
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 2

Figure 2. Superresolution of Bax by single molecule localization microscopy reveals non-random structures in apoptotic mitochondria

Overview of a reconstructed superresolution image of GFP-Bax overexpressing HeLa cells stained with AF647-anti-GFP-nanobodies. Image was acquired on fixed cells 3 h after apoptosis induction with STS. Dotted line shows the cell shape (see Fig EV3). Scale bar, 5 µm.

Magnified reconstructed superresolution image corresponding to the white rectangle in A showing the shapes of GFP-Bax WT structures (white arrows). Scale bar, 500 nm.

Gallery of typical GFP-Bax WT structures during apoptosis found in all the analyzed cells. Scale bars, 100 nm.

Overview (left) and 3 enlarged insets (right) of HeLa cells overexpressing GFP-Bax 1-2/L-6 stained with AF647-anti-GFP-nanobodies. Figure shows reconstructed superresolution images acquired without apoptosis induction. Scale bars, 5 µm (overview) and 500 nm (insets).
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 3

Figure 3. Bax wild type adopts different structures compared to the inactive mutant Bax 1-2/L-6.

Quantification of the structures found on Bax wild type (purple) and on Bax 1-2/L-6 (grey) overexpressing cells. Data show the total number of structures in all the measurements. n (Bax wild type)= 13, n (Bax 1-2/L-6)=11.

Cumulative distribution plots of GFP-Bax wild type (purple) and GFP-Bax 1-2/L-6 (grey) showing differences in amplitude and frequency. Each plot represents a single cell. Thick lines represent average cumulative distributions for all the single cells transfected with GFP-Bax WT or mutant GFP-Bax 1-2/L-6.
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 4

Figure 4. Bax non-random structures shown on HCT116 Bax/Bak -/- cells by SMLM and on HeLa cells by STED microscopy

Overview of a reconstructed superresolution image of GFP-Bax overexpressing HCT116 Bax/Bak -/- cells stained with AF647-anti-GFP-nanobodies Image was acquired on fixed cells 7 h after apoptosis induction with STS. Dotted line shows the cell shape (see Fig EV3). Scale bar, 5 µm.

Gallery of typical GFP-Bax WT non-random structures during apoptosis on HCT116 Bax/Bak -/- cells. Scale bars, 100 nm.

Comparison of confocal (left) and STED (right) images of GFP-Bax overexpressing HeLa cells 2h after STS treatment. STED reveals a notable increase in resolution. Scale bars, 500 nm.

Representative GFP-Bax non-random structures found with STED (lines, arcs, rings, double lines). Scale bars, 100 nm.
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 5

Figure 5. Transmission electron microscopy reveals mitochondrial outer membrane pores on apoptotic cells.

Representative TEM pictures of HeLa wild-type cells. Bottom panels show healthy mitochondria. Scale bars, 500nm (overview) and 200 nm (zooms).

Representative TEM pictures of apoptotic HeLa wild-type cells 4h after STS treatment (1µM). Black arrowheads point to defects on the mitochondrial outer membrane. Scale bars, 500nm (overview) and 200 nm (zooms)

Representative TEM pictures of apoptotic HeLa GFP-Bax transfected cells 4h after STS treatment (1µM). Black arrowheads point to defects on the mitochondrial outer membrane. Scale bars, 500nm (overview) and 200 nm (zooms).
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 6

Figure 6. Bax localizes heterogeneously to discrete mitochondrial foci during apoptosis.

Left panel shows the individual channels and overlaid images of confocal live cell imaging of HeLa cells overexpressing GFP-Bax (green) and mito-dsRed (magenta) at 0 and 85 minutes after apoptosis induction with STS 1µM. Right panel shows zoomed overlaid images of HeLa cells following GFP-Bax translocation (green) and mitochondrial fragmentation (magenta) during apoptosis induction with STS 1µM. Scale bars, 10 µm (left) and 2 µm (right)

Reconstructed dual-color superresolution image of GFP-Bax overexpressing HeLa cells. GFP-Bax was imaged by immunosstaining with AF647-anti-GFP-nanobodies and mitochondria were immunostained with CoxIV (primary) and cf680 (secondary) antibodies. Image was acquired 3h after STS treatment. Left panel shows the cell overview (scale bar, 5 µm), right panel up shows the overview inset (scale bar, 500nm) and right panel down shows a non-random linear GFP-Bax assembly at mitochondria (scale bar, 100nm).

(C) Reconstructed dual-color superresolution image of HeLa cells co-overexpressing mMaple-mito and GFP-Bax and stained with AF647-anti-GFP-nanobodies. Image was acquired 3h after STS treatment. Lower panel shows enlarged insets corresponding to the white rectangles in the upper panel. Scale bars, 1 µm (overview) and 100 nm (insets)
Bax assembly into rings and arcs in apoptotic mitochondria is linked to membrane pores (2015)
Figure 7

Figure 7. Full rings and arc-shaped assemblies of Bax are associated with membrane pores, as visualized by AFM.

AFM image of a control SLB prepared from LUVs with a mitochondrial-like lipid composition. The bilayer looks flat, without aggregates or defects. Scale bar, 1 µm.

AFM image of a SLB prepared from LUVs with a mitochondrial-like lipid composition pre-incubated with heat-activated Bax. The green arrows indicate the presence of membrane pores, which are heterogeneous in size and shape. The edges of the pores present protrusions corresponding to Bax clusters. Scale bar, 1 µm.

3D AFM topography of a Bax arc (left) and ring (right). Both images reveal a circular dark hole that spans the lipid membrane (green). Bax molecules around the pore rim (magenta and white) protrude 3,97 ± 1,02 nm above the membrane plane, as confirmed by the height profiles shown below each image (corresponding to the white line in the 2D image insets). The topography of the arc structure reveals a pore only partially surrounded by Bax molecules, while lipids alone form the rest of the pore rim. Images are shown in a 42° tilted representation.

Quantitative analysis of the distinct structures found for activated Bax on SLBs. Data show the total number of structures in all the measurements.

If not differently specified, the scale bar in all the images is 100 nm.
Absolute quantification of translational regulation and burden using combined sequencing approaches (2019)
Figure 3

Figure 3: Simultaneous quantification of transcription and translation of endogenous genes and a synthetic genetic construct. (A) Comparison of protein synthesis rate of endogenous E. coli genes measured using Ribo-seq from this study (in molecules/s units) and from that by Li et al. (Li et al, 2014) (in molecules/generation units). Each point corresponds to a single gene and color denotes the ratio of transcription initiation rate to translation initiation rate (giving RNAP/ribosome) capturing whether transcription (light yellow) or translation (dark blue) is more dominant. (B) Transcription (bottom) and translation (top) profiles for yggN, rpoH and greA, computed from the RNA-seq and Ribo-seq data without induction. Positions of the genetic parts and gene are shown below the profiles. (C) Promoter strengths in RNAP/s units and RBS initiation rates in ribosome/s units. (D) Transcription (bottom) and translation (top) profiles for lacZ. Profiles are shown for cells in the absence and presence of IPTG (1 mM). Position of genetic parts and gene is shown below the profiles. RBS is omitted from the genetic design due to its size. (E) Measured promoter strength in RNAP/s units, RBS initiation rate in ribosomes/s units, and the transcriptional terminator and translation termination efficiency for lacZ. Data shown for cells in the absence and presence of IPTG (1 mM).
Microbiota Dysbiosis influences Immune System and Muscle Pathophysiology of Dystrophin Deficient Mice (2022)
Figure 5

Figure 5. Microbiota depletion induces modulation of immune cells

(A-D) FACS analysis of spleen and muscle homogenates from 3m C57Bl (n=4), mdx (n=5) and mdx+ABX (n=7) mice demonstrates no significant alteration of CD45+CD11b+CD4-CD8- myeloid cells (A and C) and few differences in CD4+ or CD8+ naïve (CD62L+ CD44-), central memory (CD62L+ CD44+) and effector (CD62L- CD44+) T cells (B and D).

(E) FACS analysis of spleen of 3m C57Bl (n=7), mdx (n=9) and GFmdx (n=5) mice revealed similar proportions of CD4+ and CD8+ T cells but reduced activated CD44+ T cells in GFmdx mice. Representative plots are depicted.

(F) Graphs show cumulative frequencies of CD4+ and CD8+ T cells on live cells of 3m C57Bl (n=7), mdx (n=4) and GFmdx (n=5) mice. Representative dot plots and cumulative frequencies of splenic CD4+GITR+CD25+ Treg. Frequencies of effector CD44+ T cells were significantly decreased in spleen of GFmdx mice.

(G) Representative dot-plots showing the proportion of muscle-infiltrating CD45+ cells of 3m C57Bl (n=6), mdx (n=6) and GFmdx (n=5) mice. Cumulative frequencies of muscle-infiltrating CD45+ cells are shown.

(H) Representative images of TA muscles from 3m mdx and GFmdx mice stained for CD45 (in green), isolectin (in red) and phalloidin (in purple). Nuclei were counterstained with DAPI (in blue). Scale bar: 10 μm.

(I) Absolute number of CD3+ inflammatory cells (white arrows) were quantified in n=12 images of TA of 3m C57Bl, 3m mdx and 3m GFmdx mice (n=6 each). CD3 staining is shown in green and DAPI in blue. Scale bars: 50 μm.

Data information: Data are presented as mean ± SD. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001 ordinary one-way ANOVA, Tukey's multiple comparison test. The comparisons among the averages of CD3+ cells were evaluated using unpaired t-test.
Microbiota Dysbiosis influences Immune System and Muscle Pathophysiology of Dystrophin Deficient Mice (2022)
Figure 6

Figure 6. Muscle homeostasis of 3m mdx mice is influenced by microbiota

(A, B) RNA datasets clustering (A) and convergently up and downregulated genes (B) of muscles of 3m mdx (n=3), mdx+ABX (n=3) and GFmdx (n=3) mice.

(C) Gene onthology (GO) analysis on both groups of convergent genes.

(D) RT-qPCR analysis of TA muscles of two independent experiments with 3m mdx (n=4), mdx+ABX (n=4) and GFmdx (n=5) mice determined the expression of myogenic markers.

(E) Representative Gomori modified staining and quantification of myofiber area and relative frequency of the myofiber cross-sectional area (CSA) expressed as the frequency distribution of the TA muscles of 3m C57Bl (n=4), 3m mdx (n=4), mdx+ABX (n=4) and GFmdx mice (n=5). Pooled samples for each group with n=6240 for 3m C57Bl; n=6001 for 3m mdx; n=10556 for 3m mdx+ABX; n=23059 for GFmdx. For morphometric analysis, images were quantified with Image J software for each mouse. Scale bars: 50 μm.

(F) Quantification of the fibrotic area from Gomori stained images (pooled samples for each group with n=223 for 3m C57Bl, 3m mdx and 3m mdx+ABX; n=215 for GFmdx) and RT-qPCR analysis of Col1a (two independent experiments with n=4 animals each group).

(G) Representative images of skeletal muscle showed the distribution and composition of the myosin heavy chain (MyHC) isoforms (Type IIa, Type IIx and Type IIb).

(H, I) Graph portrays (H) the percentage of myofibers expressing different MyHC isoforms and (I) myofibers area per type of MyHC in TAs of 3m mdx (n=4), mdx+ABX (n=4) and GFmdx (n=5) mice (n=12 images per animal).

(J) Representative SDH staining and quantification of percentage of SDH+ myofibers of TAs from 3m mdx (n=4), mdx+ABX (n=4) and GFmdx mice (n=5) (n=12 images per animal). Scale bars: 50 μm.

(K) Tetanic force of TA muscle of 3m C57Bl (n=4), mdx (n=4), mdx+ABX (n=4) and GFmdx (n=5) mice.

(L) ALT, AST and CPK serum levels were measured in 3m mdx (n=4), mdx+ABX (n=4) and GFmdx mice (n=5) (two independent experiments).

Data information: Data are presented as mean ± SD (*p<0.05, **p<0.01, ***p<0.001; ****p<0.0001, ordinary one-way ANOVA, Tuckey multiple comparison test for WB and non-parametric test followed by Kruskal-Wallis test for RT-qPCR).
Microbiota Dysbiosis influences Immune System and Muscle Pathophysiology of Dystrophin Deficient Mice (2022)
Figure 7

Figure 7. Effects of dysbiotic microbiota of mdx on intestinal, spleen and muscle inflammation and muscle function.

(A) FACS analysis of T cell subsets from lamina propria in 3m C57Bl (n=5), mdx (n=5) and ABX-mdxFMT_C57Bl (n=5/6) showing decrease in CD3+T cells in ABX-mdxFMT_C57Bl. Infiltrating CD3+CD4+, and regulatory CD69+ subsets of CD4+ and CD8+ were decreased in ABX-mdxFMT_C57Bl. Eubiotic FMT in mdx modulates T helper response, with reductions in the cumulative frequencies of CD4+ IFNγ+ (Th1) and CD4+ IL-10+ cells in ABX-mdxFMT_C57Bl. Data are presented as mean ± SD (*p<0.05; **p<0.01ordinary one-way ANOVA, Tukey's multiple comparison test.

(B-C) FACS analysis of spleen and muscle homogenates from 3m C57Bl (n=5), mdx (n=5) and ABX-mdxFMT_C57Bl (n=5/6). Analysis of spleen revealed downregulation of Ly6C+ inflammatory monocytes and F4/80+ macrophages in ABX-mdxFMT_C57Bl. Eubiotic FMT in mdx mice determined a decrease of CD4+/CD8+ CD44+CD62L effector and GITR+CD4+ T-cells in ABX-mdxFMT_C57Bl. Gut-derived CCR9+CD8+TEM+ cells were increased in ABX-mdxFMT_C57Bl. Data are presented as mean ± SD (*p<0.05; **p<0.01; ***p<0.001; ****p<0.0001, ordinary one-way ANOVA, Tukey's multiple comparison test.

(C) Graphs showing cumulative frequencies of infiltrating CD45+CD4+ and CD45+CD8+ cells in muscles of 3m C57Bl (n=5), mdx (n=5) and ABX-mdxFMT_C57Bl (n=5/6) were decreased in ABX-mdxFMT_C57Bl related to mdx mice. . Data are presented as mean ± SD (*p<0.05; **p<0.01, ordinary one-way ANOVA, Tukey's multiple comparison test.

(D, E) Representative H&E staining (D) and quantification of myofiber area with Image J software (E) of TA muscles from 3m C57Bl (n=5), mdx (n=5) and ABX-mdxFMT_C57Bl (n=6). Scale bars for H&E: 200 μm.

(F) Measurement of ALT, and CPK in the serum of 3m C57Bl (n=5), mdx (n=5) and ABX-mdxFMT_C57Bl (n=6).

(G) Tetanic force of TA muscles from mdx (n=5) and ABX-mdxFMT_C57Bl (n=6).

(H) Representative images of skeletal muscle showed the distribution and composition of MyHC isoforms (Type IIa, Type IIx and Type IIb). Scale bar: 50 μm.

(I) Graph portrays the percentage of myofibers expressing different MyHC isoforms. n=12 images were analysed for each mouse.

(J-K) Representative SDH staining and quantification of percentage of SDH+ myofibers of TA muscles from mdx (n=5) and ABX-mdxFMT_C57Bl (n=6) (n=12 images per mouse). Scale bar: 200 μm.

(L-M) Representative image of CD31 (in cyan), α-SMA (in green) and isolectin (in red) staining and their quantification in TA muscles from mdx (n=5) and ABX-mdxFMT_C57Bl (n=6) mice. Scale bar: 500 μm.

Data information: Data for tetanic force, ALT and CPK concentration and staining quantification are presented as mean ± SD (*p<0.05, **p<0.01, ***p<0.001; ****p<0.0001, one-way ANOVA, Tukey's multiple comparison test).

Supplementary Figure Legends
The TOR-dependent phosphoproteome and regulation of cellular protein synthesis (2021)
Figure 1

Figure 1 - TOR activity is a major regulator of cellular protein synthesis rates

Measuring active protein synthesis using the methionine analogue, L-homopropargylglycine (HPG), and measuring the change in rates of mass doubling and protein synthesis upon inhibition of TOR activity.

A Density plot showing the measured fluorescence intensity distribution of individual cells for each timepoint after HPG addition. Time is represented on the y-axis with the bottom of the distribution plot aligning to the corresponding timepoint.

B Median fluorescence intensity of the distribution plots in A plotted against the indicated time after HPG addition. Linear regression line fitted from 5 minutes onwards.

C, D Growth rates as measured by the change in optical density (OD595) over time for wild type (C) and Torin1-resistant (D) cells upon 5 µM of Torin1 treatment (at T = 0 min). Respective strains were treated with the DMSO vehicle alone to control for potential effects of the solvent on cell growth. Logarithmic values of the OD595 increase are plotted against time in minutes. Data aggregated from 3 independent experiments with error bars representing the standard deviations (SD). Linear regressions were fitted using the curve fitting function on Prism9. Ratios in the rate of mass doubling (as calculated from gradient of regression lines detailed in Methods) of Torin1 treated cells to DMSO control were 0.34 in wild type (C) and 0.97 for Torin1-resistant (D) cells.

E Rate of protein synthesis as measured by the amount of HPG incorporation after 20 mins of differing time intervals (0 - 60 mins) of 5 µM Torin1 for wild type (green) and tor1∆ (yellow). Values are normalised to T = 0 minutes. Exponential one-phase decay non-linear regressions were fitted using the curve fitting function on Prism9. Data aggregated from 3 independent experiments with error bars representing the standard deviations (SD).

F Growth rates for wild type and tor1∆, as measured by the change in optical density (OD595) over time. Linear regressions fitted with Prism9.
The TOR-dependent phosphoproteome and regulation of cellular protein synthesis (2021)
Figure 2

Figure 2 - TOR-dependent phosphoproteomic changes identifies conserved proteins with potential roles in modulating rates of protein synthesis

Investigating the protein expression levels for the phosphoproteomics time course study and validation of experimental conditions based on known indirect TOR phosphorylation targets.

A Scatter plot showing the protein levels in wild type cells after 40 minutes of Torin1 (5 µM) addition plotted against the corresponding values for the same protein in Torin1-resistant cells after 40 minutes. Dark grey dashed lines at x = ±1 indicate the 2-fold threshold, of which no proteins exceeded in the study. The light grey dashed line represents y = x, plotted as guide to indicate positive correlation in protein levels between wild type and Torin1-resistant cells.

B Frequency distribution of the proteins levels after 40 minutes of Torin1 treatment in wild type cells (n = 3380), with 98% of all proteins (3314/3380) within a 1.3-fold boundary, as indicated by the grey dashed lines at ±0.38.

C - F Phosphorylation changes of the conserved phosphosites, S235 and S236, on Rps601(C) and Rps602 (D - F), over 85 minutes of Torin1 (5 µM) treatment wild type (green) and Torin1-resistant (brown) cells. M-values (in brackets) represent the different multiplicities of the same phosphosites. Lines were fitted with the curve fitting function on Prism9 using the the non-linear regressions exponential one-phase decay (C, D) or plateau followed by one-phase decay (E, F) functions respectively. Grey dashed lines indicate 2-fold threshold.