Fig. 2: NPM1 loss suppresses WNT-driven hyperproliferation and tumor formation in vivo.
From: Nucleophosmin supports WNT-driven hyperproliferation and tumor initiation
a, Npm1 relative expression in SI and colon tissues from WT mice and a range of genetically engineered murine models of CRC (GSE309379; n = 5: WT (SI and colon), VilCreERT2BrafV600E/+ (colon), VilCreERT2BrafV600E/+Trp53loxP/loxPALK5loxP/loxPR26LSL-N1icd/+ (SI); n = 4: VilCreERT2 (SI and colon), VilCreERT2BrafV600E/+Trp53loxP/loxPALK5loxP/loxP (SI), VilCreERT2ApcloxP/loxPKRasG12D/+ (colon), VilCreERT2ApcloxP/loxPKRasG12D/+Trp53loxP/loxP (SI), VilCreERT2ApcloxP/loxPTrp53loxP/loxP (SI), AhCreERT2ApcloxP/loxPMycloxP/loxP (SI); n = 3: VilCreERT2KRasG12D/+Trp53loxP/loxP (SI), VilCreERT2ApcloxP/loxPKRasG12D/+Trp53loxP/loxP (colon); n = 7: VilCreERT2BrafV600E/+Trp53loxP/loxP (SI and colon), VilCreERT2KRasG12D/+Trp53loxP/loxP R26LSL-N1icd/+ (SI); n = 10: VilCreERT2ApcloxP/loxP (SI); n = 12: VilCreERT2ApcloxP/loxP (colon); n = 8: VilCreERT2ApcloxP/loxPKRasG12D/+ (SI); n = 6: AhCreERT2ApcloxP/loxP (SI)). Data were statistically assessed by one-way analysis of variance (ANOVA) followed by Dunnett’s multiple comparisons test. b, Representative micrographs from ApcloxP/loxPNpm1+/+ (n = 5) and ApcloxP/loxPNpm1loxP/loxP (n = 5) SI sections stained with hematoxylin and eosin (H&E) and anti-bromodeoxyuridine (BrdU) from mice collected 4 days after induction. The red bars indicate the expanded crypt depth after APC loss. c, Quantification of BrdU+ cells in SI half-crypts of animals from the groups shown in a (n = 5 per group). Data were statistically assessed using an unpaired, two-tailed Mann–Whitney U-test. d, Survival curves of ApcloxP/+ (n = 14) and ApcloxP/+Npm1loxP/loxP (n = 16) mice sampled at the clinical endpoint. Median survival in days is indicated in parentheses. Censored mice are denoted as tick marks at the indicated times after induction. The P value was obtained using a log-rank (Mantel–Cox) test. e, Tumor numbers from ApcloxP/+ (n = 8) and ApcloxP/+Npm1loxP/loxP (n = 9) mice sampled at the clinical endpoint. Data were statistically assessed using an unpaired, two-tailed t-test. f, Representative staining for NPM1 on SI tissue sections from ApcloxP/+Npm1loxP/loxP animals at the clinical endpoint (n = 5). g, Quantification of the percentage of tumors being positive, negative or mosaic for NPM1 expression in each ApcloxP/+Npm1loxP/loxP animal at the clinical endpoint (n = 5). h, Survival curves of Lgr5-CreERT2ApcloxP/loxP (n = 13) and Lgr5-CreERT2ApcloxP/loxPNpm1loxP/loxP (n = 11) mice sampled at the clinical endpoint. Median survival in days is indicated in parentheses. Censored mice are denoted as tick marks at the indicated times after induction. The P value was obtained using a log-rank (Mantel–Cox) test. i, Tumor numbers from Lgr5-CreERT2ApcloxP/loxP (n = 11) and Lgr5-CreERT2ApcloxP/loxPNpm1loxP/loxP (n = 9) mice sampled at the clinical endpoint. Data were statistically assessed using an unpaired, two-tailed Mann–Whitney U-test. j, Representative staining for NPM1 on SI tissue sections from Lgr5-CreERT2ApcloxP/loxP (n = 3) and Lgr5-CreERT2ApcloxP/loxPNpm1loxP/loxP (n = 7) animals at the clinical endpoint. k, Quantification of the percentage of tumors being positive, negative or mosaic for NPM1 expression in each Lgr5-CreERT2ApcloxP/loxPNpm1loxP/loxP animal at the clinical endpoint (n = 7). l, Representative micrographs from ApcloxP/loxPKrasG12D/+Npm1+/+ (n = 4) and ApcloxP/loxPKrasG12D/+Npm1loxP/loxP (n = 4) SI sections stained with H&E and anti-BrdU from mice collected 3 days after tamoxifen induction. The red bars indicate the expanded crypt depth after APC loss and KRASG12D activation. m, Quantification of BrdU+ cells in the SI of animals from the groups shown in l (n = 4 per group). Data were statistically assessed using an unpaired, two-tailed t-test. n, Survival curves of ApcloxP/+KrasG12D/+ (n = 17) and ApcloxP/+KrasG12D/+Npm1loxP/loxP (n = 20) mice sampled at the clinical endpoint. Median survival in days is indicated in parentheses. The P value was obtained using a log-rank (Mantel–Cox) test. o, Tumor numbers from ApcloxP/+KrasG12D/+ (n = 17) and ApcloxP/+KrasG12D/+Npm1loxP/loxP (n = 20) mice sampled at the clinical endpoint. Data were statistically assessed using an unpaired, two-tailed Mann–Whitney U-test. p, Representative staining for NPM1 on colon tissue sections from ApcloxP/+KrasG12D/+ and ApcloxP/+KrasG12D/+Npm1loxP/loxP animals at the clinical endpoint. q, Quantification of the percentage of tumors being positive, negative or mosaic for NPM1 expression in each ApcloxP/+KrasG12D/+Npm1loxP/loxP animal at the clinical endpoint (n = 9) compared to that of ApcloxP/+Npm1loxP/loxP animals (n = 5) presented in Fig. 2g. Data were statistically assessed using a two-way ANOVA with Šidák’s correction for multiple comparisons. The bar charts present data as the mean ± s.e.m.; the boxes in the box plots extend from the 25th to 75th percentile, the whiskers extend to the minimum and maximum values, and the line in every box is plotted at the median. Statistically significant P values are shown in red. b,l, Scale bar, 50 μm. f, Scale bar, 1 mm. j, Scale bar, 300 μm. p, Scale bar, 500 μm.
