Down syndrome (DS) patients suffer from mental retardation, but also display enhanced beta-APP production and develop cortical amyloid plaques at an early age. As beta-APP and Notch are both processed by gamma-secretase, we analyzed expression of the Notch signaling pathway in the adult DS brain and in a model system for DS, human trisomy 21 fibroblasts by quantitative PCR. In adult DS cortex we found that Notch1, Dll1 and Hes1 expression is up-regulated. Moreover, DS fibroblasts and Alzheimer disease cortex also show overexpression of Notch1 and Dll1, indicating that enhanced beta-APP processing found in both DS and AD could be instrumental in these changes. Using pull-down studies we could demonstrate interaction of APP with Notch1, suggesting that these transmembrane proteins form heterodimers, but independent of gamma-secretase. We could demonstrate binding of the intracellular domain of Notch1 to the APP adaptor protein Fe65. Furthermore, activated Notch1 can trans-activate an APP target gene, Kai1, and vice versa, activated APP can trans-activate the classical Notch target gene Hes1. These data suggest that Notch expression is activated in Down syndrome, possibly through cross-talk with APP signaling. This interaction might affect brain development, since the Notch pathway plays a pivotal role in neuron-glia differentiation.