DLL1 (top) studs the surface of the cell, whereas DLL3 (bottom) collects in the interior.

Two key developmental proteins that once appeared to be molecular twins perform different jobs and work in different parts of the cell, as Geffers et al. show on page 465.

Among its many functions, the Notch pathway helps orchestrate the formation of somites that give rise to the vertebral column and other structures. The pathway appears to be full of redundancy. For example, mice carry four Notch genes and three for the Delta proteins that interact with the pathway. The jobs of two of the Delta proteins, DLL1 and DLL3, have remained unclear. Initial studies suggested that the proteins are interchangeable. But individually deleting the proteins, which are made in the same part of the mesoderm, induces different developmental defects, and a recent study suggested that DLL3 interferes with DLL1.

To sort through the confusion, Geffers et al. replaced one or both copies of the Dll1 gene with a short but functional version of the Dll3 gene. Mice lacking DLL1 die as embryos. The researchers found that mice that manufacture DLL3 instead of DLL1 also perished during embryogenesis, indicating that one protein cannot substitute for the other.

The team also gauged whether one protein inhibits the activity of the other by examining mice with three copies of Dll3 and one of Dll1. Animals with only one copy of Dll1 display some skeletal flaws. If the proteins were antagonists, the scientists reasoned, animals with the extra DLL3 should show even more severe abnormalities. But these mice were no worse off.

The researchers confirmed their findings by tracking Notch after its activation. When Notch switches on, its intracellular tail detaches and spirits the signal to the nucleus. Levels of this tail piece did not increase in embryos lacking DLL3, the scientists determined. DLL3 thus does not interfere with Notch activation.

So what is DLL3 doing? The researchers are still not sure, but they discovered that most of the protein lurked inside the cell, mainly in the Golgi apparatus, rather than on the surface, where DLL1 resided. The Golgi location suggests that, instead of triggering Notch, DLL3 has some other, undiscovered role in the pathway.