Fig. 4

mTOR and p38 activity in relation to cell size. A Representative image of hematopoietic stem cells from wild-type mice stained for Phospho-S6 (P-S6, Alexa 488) and DNA (DAPI). B Values obtained from Cell-ACDC segmentation using Cellpose on bright-field signal: Alexa 488 total fluorescence intensity per cell volume (a.u., proxy for mTOR activity) of phospho-S6 ribosomal protein (Ser240/244) staining as a function of the HSC volume (fL). Gates of XS-, M- and XL-sized HSCs are indicated in grey (n=1626 cells). Alexa 488 total fluorescence intensity per cell volume (a.u.) of phospho-S6 ribosomal protein (Ser240/244) staining in control (no primary antibody), all HSCs, XS-sized HSCs (n=247), M-sized HSCs (n=493), and XL-sized HSCs (n=244). C Representative image of hematopoietic stem cells from wild-type mice stained for Phospho-p38 (P-p38, Alexa 488) and DNA (DAPI). D Values obtained from Cell-ACDC segmentation (using StarDist on DAPI signal): Alexa 488 total fluorescence intensity per nuclear volume (a.u., proxy for p38 activity) of Phospho-p38 mitogen-activate protein kinase (MAPK, Thr180/Tyr182) staining as a function of the HSC nuclear volume (fL). Gates of XS-, M- and XL-sized HSCs are indicated in grey (n = 586). Alexa 488 total fluorescence intensity per cell volume (a.u.) of phospho-p38 (Thr180/Tyr182) staining in control (no primary antibody, n = 196), all HSCs, XS-sized HSCs (n = 90), M-sized HSCs (n = 179), and XL-sized HSCs (n = 77)