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. 2023 Jan 19;55(2):232–245. doi: 10.1038/s41588-022-01280-z

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 8 — a, Notch1^+/+ and Notch1^−/− normal esophageal tissue and tumors (Fig. 7a) were RNA sequenced. Notch1^+/+: n = 11 epithelial samples from seven mice, n = 8 Notch1^+/+ tumors from four mice; Notch1^−/− n = 10 epithelial samples from seven mice, n = 6 Notch1^−/− tumors from five mice. b, MA plots showing differentially expressed genes (red, q < 0.05, DESeq2 analysis, two-sided Wald test with Benjamini–Hochberg correction), red, in Notch1^+/+ and Notch1^−/− tumors versus normal epithelium. Zero-fold change shown by red dotted line. c, −log₁₀ (P value) of top Gene ontology biological processes (GOBP) in tumors versus normal epithelium in Notch1^+/+, gray, Notch1^−/−, red genotypes (Supplementary Tables 27 and 28). d, MA plots showing differentially expressed genes (red, q < 0.05, DESeq2 analysis, two-sided Wald test with Benjamini−Hochberg correction), in tumors from Notch1^−/− versus Notch1^+/+ esophagus. Red dotted line, zero-fold change. e, GSEA of tumors from Notch1^−/− versus Notch1^+/+- esophagus, DNA replication gene set shown (normalized enrichment score, NES = −2.48, false discovery rate, FDR q-value = 0.0, Supplementary Table 29). f, Transcript per million values of cell cycle and DNA replication transcripts selected from GSEA in tumors from Notch1^+/+and Notch1^−/− esophagus. Mean ± s.e.m., n = 8 tumors from Notch1^+/+ esophagus and n = 6 from Notch1^−/− esophagus. Two-tailed unpaired Student’s t-test. g, Representative images of n = 8 tumors from Notch1^+/+ and n = 9 tumors from Notch1^−/− esophagus. KRT14 (green), pHH3 (gray). DNA, blue. Scale bars, 30 µm. h, Percentage of pHH3⁺, KRT14⁺ keratinocytes within tumors from Notch1^+/+ and Notch1^−/− esophagus. Mean ± s.e.m., each dot represents a tumor, +/+: n = 8 tumors from 4 mice; −/−: n = 9 tumors from 7 mice. Two-tailed unpaired Student’s t-test. i, Representative images from n = 8 tumors each from Notch1^+/+ and Notch1^−/− esophagi, KRT14 (green), phospho-ERK1/ERK2 (p-ERK, magenta), DNA (blue). Insets, magnified areas indicated by white squares. Scale bars, 30 µm. j, Normalized mean intensity of fluorescence for p-ERK (left) and total ERK (t-ERK, right) in KRT14⁺ cells in tumors from Notch1^+/+ and Notch1^−/− esophagi relative to adjacent normal tissue. Mean ± s.e.m., each dot represents a tumor. +/+: n = 8 tumors from four mice; −/−: n = 8 tumors from seven mice. Two-tailed unpaired Student’s t test. k, In tumors lacking Notch1, signals downstream of mutant Atp2a2 are disrupted, cell division reduced, and tumor growth slows. A.U., arbitrary unit. See Supplementary Tables 24 and 27–30.