Fig. 6: The AIRIMV190G cerebral organoids exhibit impaired radial glial cell fate specification.
From: A programmed decline in ribosome levels governs human early neurodevelopment
a, UMAP showing FABP7 expression in control and AIRIM variant cells in day 15 organoids. b, IF image of day 15 control and mutant (V190G and R72W) organoids of transitioning NE marker ZEB2 and committed RG marker BLBP (encoded by FABP7). Note that the mutant exhibits less expression of BLBP and less-organized nuclei (marked by ZEB2). Scale bar, 20 µm. c, Quantification of mean BLBP IF intensity of day 15 control and mutant (V190G) organoids. *P = 0.0463. Unpaired two-tailed t-test with Welch’s correction. WT, n = 7; V190G, n = 8 imaged regions from two independent batches. Error bars show s.e.m. d, Quantification of mean BLBP IF intensity of day 15 control and mutant (R72W) organoids. ****P = 8.65486 × 10−5. Unpaired two-tailed t-test with Welch’s correction. WT, n = 8; R72W, n = 8 imaged regions from two independent batches. Error bars show s.e.m. e, IF image of day 15 control and AIRIMV190G, AIRIMR72W and AFG2BI466M/V245E organoids of the progenitor marker Sox2 and the neuronal differentiation marker TUBB3. Scale bar, 20 μm. Also included are IF images of day 30 control and AIRIMR72W organoids stained for the progenitor marker Sox2 and the neuronal differentiation marker TUBB3. Scale bar, 40 μm. Three independent batches were performed. f, Representative whole-mount organoid IF images showing the morphology of neural progenitor cells (PAX6+), around apical (ZO1+) lumens, revealed by sparse labelling with GFP in control and mutant (V190G) organoids. Day 10 cells are columnar and exhibit typical NE morphology. Day 15 control cells show a thinning of apical processes (yellow arrows), whereas mutant cells still seem to be columnar. Scale bar, 20 μm. g, Quantification of the length of neural progenitor cells in control and mutant (V190G) organoids at days 10, 13 and 15, showing reduced length of the progenitor cells in mutant compared with control organoids. Cells with clear apical and basal labelling were used for quantification. Mann–Whitney U-test, ****P < 0.0001, two-tailed, multiple-comparison corrected. Day 10 control, n = 10 cells; day 10 V190G, n = 8 cells; day 13 control, n = 14 cells; day 13 V190G, n = 16 cells; day 15 control, n = 8 cells; and day 15 V190G, n = 8 cells. Padj = 0.0714, day 10; Padj = 9.177792 × 10−7, day 13; Padj = 1.098352 × 10−7, day 15. Error bars show s.e.m. h, Representative whole-mount organoid IF images showing the morphology of neural progenitor cells revealed by sparse labelling with GFP in control and mutant (R72W) organoids. Day 10 cells are columnar and exhibit typical NE morphology. Day 15 control cells show a thinning of apical processes, whereas mutant cells do not. Scale bar, 20 μm. i, Quantification of the length of neural progenitor cells in control and AIRIMR72W organoids at days 10 and 15. Cells with clear apical and basal labelling were used for quantification. Mann–Whitney U-test, ***P = 0.0009, two-tailed, day 10 control, n = 12 cells; day 10 V190G, n = 9 cells; day 15 control, n = 11 cells; and day 15 V190G, n = 10 cells. Error bars show s.e.m.
