The Notch signalling pathway has a crucial function in determining cell fates in 
multiple tissues within metazoan organisms. On binding to ligands, the Notch 
receptor is cleaved proteolytically and releases its intracellular domain 
(NotchICD). The NotchICD enters the nucleus and acts cooperatively with other 
factors to stimulate the transcription of target genes. High levels of 
Notch-mediated transcriptional activation require the formation of a ternary 
complex consisting of NotchICD, CSL (CBF-1, suppressor of hairless, LAG-1) and a 
Mastermind family member. However, it is still not clear how the formation of 
the ternary complex is regulated. Here we show that Nemo-like kinase (NLK) 
negatively regulates Notch-dependent transcriptional activation by decreasing 
the formation of this ternary complex. Using a biochemical screen, we identified 
Notch as a new substrate of NLK. NLK-phosphorylated Notch1ICD is impaired in its 
ability to form a transcriptionally active ternary complex. Furthermore, 
knockdown of NLK leads to hyperactivation of Notch signalling and consequently 
decreases neurogenesis in zebrafish. Our results both define a new function for 
NLK and reveal a previously unidentified mode of regulation in the Notch 
signalling pathway.