Extended Data Figure 9. Secondary validation that asparagine availability regulates EMT.

a, Transcription-level changes in EMT-up and -down genes that occur in response to Asns silencing in 4T1-T cells (n = 2 replicates per condition, edges of the box are the 25th and 75th percentiles and error bars extend to the values q3 + w(q3 − q1) and q1 − w(q3 − q1), in which w is 1.5 and q1 and q3 are the 25th and 75th percentiles, which is also the case for b–e, h and i, rank-sum P < 0.001 for EMT-up genes, DESeq false discovery rate < 0.05 for Twist1 and Cdh1). b, Transcription-level changes in EMT-up and -down genes that occur in response to the medium of Asns-silenced 4T1-T cells being supplemented with l-asparagine (n = 2 replicates per condition, rank-sum P < 0.005 for EMT-up genes). c, Gene expression changes in EMT-up and -down genes that result from ATF4 knockout in near haploid KBM-7 chronic myelogenous leukaemia (HAP-1 cells, rank-sum P < 0.05 for EMT-up genes). d, Gene expression changes in EMT-up and -down genes, which result in the liver cells of homozygous ATF4-deleted mice when treated with l-asparaginase (rank-sum P < 0.05 for EMT-down genes in wild-type mice treated with l-asparaginase (WT+ L-asp) mice, and both EMT-up and -down genes in ATF4 mice treated with l-asparaginase (ATF4+ L-asp) mice). e, Volumes of tumours resulting from orthotopic injection of Tgf-β-silenced and -expressing 4T1-T cells (n = 10 mice per cell line). f, Percentage of Twist1-positive regions based on IHC staining of sections from tumours described in e (n = 5 tumour sections per cell line, rank-sum P < 0.01). g, Percentage of Cdh1-positive regions based on IHC staining of sections from tumours described in e (n = 5 tumour sections per cell line, rank-sum P < 0.01). h, Quantification of metastases resulting from the tumours described in e (rank-sum P < 0.05). i, Quantification of metastases resulting from intravenous injection of Tgf-β -silenced and -expressing cells (n = 10 mice per cell line, rank-sum P < 0.05). See Source Data.