Activation of the single-pass transmembrane receptor Notch by the ligands Delta or Serrate mediates various cell-fate effects, which arise from signal transduction through a conserved pathway. This begins with ligand-induced cleavage of the extracellular domain (ECD) of Notch, and ends with the regulation of target genes through the association of Notch's intracellular domain (ICD) with a transcription factor. After extracellular cleavage, a γ-secretase complex — containing Presenilin, Nicastrin and other proteins — releases the intracellular domain of Notch from the membrane. Whereas Presenilin is thought to provide the catalytic activity, Nicastrin's involvement has remained elusive. Now, three groups show that Nicastrin also has an obligate role in Notch signalling.

All three groups showed that the phenotype of nicastrin (nct)-mutant Drosophila melanogaster resembles that of Notch-mutant embryos. The groups then examined the signalling potential of different forms of Notch in nct-mutant cells. Neither wild-type Notch nor a form lacking the ECD could activate Notch-mediated signalling, whereas the ICD could, indicating that Nicastrin is required to release the ICD of Notch. Moreover, as nct-null cells produced Delta and therefore induced Notch signalling in surrounding wild-type cells, but couldn't themselves respond to Delta, this role is cell-autonomous.

Struhl's group assayed for nuclear import of the Notch ICD to show that Nicastrin is needed for the transmembrane cleavage. They used Notch chimeric receptors of full-length Notch, Notch lacking the ECD or Notch containing only the ICD, fused to a transcriptional activator. Nuclear import — as measured by reporter gene activation — occurred in wild-type embryos using all three chimaeras, but only the ICD chimaeras translocated to the nucleus in nct-null embryos.

Consistent with this, St Johnston's group used antibodies directed against both the ICD and ECD of Notch to show that, in nct−/− cells, the Notch ICD remains at the membrane, whereas the ECD is cleaved normally. Fortini's group also provided direct biochemical evidence that Nicastrin is required for the transmembrane cleavage of Notch. Nicastrin loss-of-function mutants show an inhibition of specific proteolysis of Notch that is identical to that induced by a γ-secretase inhibitor.

While these data provide irrefutable evidence that Nicastrin is required for Presenilin-dependent Notch transmembrane cleavage, we have yet to find its precise role. Clones of follicle cells in late-stage egg chambers of nct-mutant embryos had reduced levels of Presenilin protein, but not messenger RNA, as did Drosophila S2 cells in which Nicastrin expression was abolished by RNA interference. Furthermore, Presenilin failed to accumulate at the apical plasma membrane of nct−/− wing disc cells, implying that defective Presenilin transport might lead to a decrease in protein stability, or vice versa. Perhaps Presenilin can only form a stable complex which is capable of moving to the membrane when Nicastrin is present. As Nicastrin can bind Notch, it is also highly plausible that it functions to recruit the Presenilin-containing complex to Notch.