L1-CAM (L1 cell-adhesion molecule), or more simply L1, plays an important role in the progression of human carcinoma. Overexpression promotes tumour-cell invasion and motility, growth in nude mice and tumour metastasis. It is feasible that L1-dependent signalling contributes to these effects. However, little is known about its mechanism in tumour cells. We reported previously that L1 is cleaved by ADAM (a disintegrin and metalloprotease) and that the cytoplasmic part is essential for L1 function. Here we analysed more closely the role of proteolytic cleavage in L1-mediated nuclear signalling. Using OVMz carcinoma cells and L1-transfected cells as a model, we found that ADAM10-mediated cleavage of L1 proceeds in lipid raft and non-raft domains. The cleavage product, L1-32, is further processed by PS (presenilin)/gamma-secretase to release L1-ICD, an L1 intracellular domain of 28 kDa. Overexpression of dominantnegative PS1 or use of a specific gamma-secretase inhibitor leads to an accumulation of L1-32. Fluorescence and biochemical analysis revealed a nuclear localization for L1-ICD. Moreover, inhibition of ADAM10 and/or gamma-secretase blocks nuclear translocation of L1-ICD and L1-dependent gene regulation. Overexpression of recombinant L1-ICD mediates gene regulation in a similar manner to full-length L1. Our results establish for the first time that regulated proteolytic processing by ADAM10 and PS/gamma-secretase is essential for the nuclear signalling of L1 in human carcinoma cell lines. Key words: a disintegrin and metalloprotease 10 (ADAM10), L1 cell-adhesion molecule (L1-CAM), nuclear translocation, presenilin (PS)/gamma-secretase activity, raft, signalling.