Notch signaling is one of key pathways constituting the stem cell signaling network. DLL1, DLL3, DLL4, JAG1 and JAG2 with DSL domain are typical Notch ligands, while DNER, F3/Contactin and NB-3 without DSL domain are atypical Notch ligands. Notch-ligand binding to NOTCH1, NOTCH2, NOTCH3 or NOTCH4 receptor induces the receptor proteolysis by metalloprotease and gamma-secretase to release Notch intracellular domain (NICD). Typical Notch ligands transduce signals to the CSL-NICD-Mastermind complex for the maintenance of stem or progenitor (transit-amplifying) cells through transcriptional activation of HES1, HES5, HES7, HEY1, HEY2 and HEYL genes, and also to the NF-kappaB-NICD complex for the augmentation of NF-kappaB signaling. Atypical Notch ligands transduce signals to the CSL-NICD-Deltex complex for the differentiation of progenitor cells through MAG transcriptional activation. Notch signals are transduced to the canonical pathway (CSL-NICD-Mastermind signaling cascade) or the non-canonical pathway (NF-kappaB-NICD and CSL-NICD-Deltex signaling cascades) based on the expression profile of Notch ligands, Notch receptors, and Notch signaling modifiers. Canonical Notch signaling is activated in the stem or progenitor domain of gastrointestinal epithelium, such as basal layer in esophagus and lower part of the crypt in colon. Notch signaling to inhibit secretory cell differentiation is oncogenic in gastric cancer and colorectal cancer, while Notch signaling to promote keratinocyte differentiation is anti-oncogenic in esophageal squamous cell carcinoma (SCC). Single nucleotide polymorphism (SNP), epigenetic change, and genetic alteration of genes encoding Notch signaling-associated molecules will be utilized as biomarkers for gastrointestinal cancer. gamma-Secretase inhibitors, functioning as Notch signaling inhibitors, will be applied as anti-cancer drugs for gastric cancer and colorectal cancer.