Fig. 5: The cytoplasm is diluted in early-differentiating cells.
From: Cell state-specific cytoplasmic density controls spindle architecture and scaling
a, Average (Avg) z-projections of 3D refractive index (RI) maps derived from optical diffraction tomography of mitotic ESCs or DIF (top). Colour-coded according to RI. Maximum-projected epi-fluorescent micrographs showing the tubulin::GFP signal (bottom). Dotted lines show cell boundaries. Representative images from n = 3 experiments. Scale bars, 5 µm. b, Cellular mass density decreases upon differentiation. 3D cellular mass densities of mitotic ESCs versus DIF. Data points show individual cells (ESCs n = 60 cells and DIF n = 70 cells pooled from three independent experiments). Boxes show interquartile ranges, black lines inside boxes denote medians, whiskers show the minima and maxima. Welch’s t-test (two-sided), P = 2.2 × 10−10. c, Average tubulin::GFP signal (3D total cell) decreases during differentiation. Data from long-term automated imaging (Fig. 1). Boxes show interquartile ranges, black lines inside boxes denote medians, whiskers show the minima and maxima. Data points show individual cells (ESCs n = 1,084 cells and DIF n = 2,920 cells pooled from nine independent experiments). Welch’s t-test (two-sided), P = 1.2 × 10−28. d, As in b but showing cellular mass density of DIF from 2i + LIF ESCs cultures (ESCs n = 52 cells and DIF n = 52 cells pooled from two independent experiments). Welch’s t-test (two-sided), P = 1.3 × 10−9. e, Spindle volume subscaling is differentiation intrinsic. Comparing spindle bulk volume scaling to cell volume in DIF originating from 2i + LIF ESCs cultures or from FBS + LIF ESCs cultures, as determined by confocal live-cell imaging (left). Data points show individual cells (ESCs (2i + LIF) n = 215, ESCs (FBS + LIF) n = 181, DIF (from 2i + LIF) n = 98, DIF (from FBS + LIF) n = 215, cells pooled from five independent experiments). Large circles represent medians in 500 µm3 cell volume bins. Error bars show interquartile ranges. Zoomed-in detail (right). ****P < 0.0001.
