Fig. 6: Growth of Ass1-deficient small intestinal organoids is impaired in arginine-low conditions only and arginine transporters could compensate for loss of Ass1.

A qRT-PCR for Ass1 in Ass1 and Apc knockout organoids. Actb and Ppia are used as reference genes. ^*p < 0.05 compared to Ass1^wt/wt Apc^wt/wt organoids, ^#p < 0.05 compared to Ass1^wt/wt Apc^−/− organoids by one-way ANOVA with Tukey’s multiple comparisons test. B Western blot for ASS1 and Vinculin in Ass1 and Apc knockout organoids. C Intracellular LC-MS measurements of M4 arginine in organoids cultured with M4 citrulline (U-13C;3,3,4-D3). D qRT-PCR for indicated urea-cycle genes. E qRT-PCR for indicated amino acid transporters. ^*p < 0.05, ^***p < 0.001, ^****p < 0.0001 by one-way ANOVA with Tukey’s multiple comparisons test. F Microscopic bright-field images of organoids cultured in regular and low-arginine medium. Representative images are shown. Scale bar represents 100 µm. G Quantification of organoid size cultured in regular and low-arginine medium (n = 3 independent experiments) ^*p < 0.05 student’s t-test.