Abstract

Neogenin-1 (NEO1) is a transmembrane receptor involved in axonal guidance, angiogenesis, neuronal cell migration, and neuronal cell death. NEO1 binds to the Netrin and RGM ligand families, however, little is known about the specific function of these proteins and their downstream signaling throughout tumor progression. Public databases contain extensive information regarding primary neuroblastoma tumors and the correlated expression of NEO1 and Netrin-4. Clinical analysis of this data reveals that high expression of both molecules in patients correlate with poor survival rate. Based on this data we sought out to determine the interaction between NEO1 and Netrin-4, and subsequently, the role of their signaling in neuroblastoma cell migration, proliferation, survival, and metastasis. Our results revealed that knocking down NEO1 reduces cell migration and that silencing of endogenous Netrin-4 reduces not only cell migration, but also proliferation and cell survival. The Overexpression of NEO1 promotes cell migration in the presence of Netrin-4, but on the other hand, it triggers apoptosis when the ligand is absent. However, addition of physiological concentrations of exogenous Netrin-4 prevented apoptosis. Metastasis analysis via in ovo chorioallantoic membrane assays of chicken embryos showed that NEO1 and endogenous Netrin-4 are involved in tumor metastasis. Finally, we found that cell migration initiated upon the binding of Netrin-4 to NEO1. When this interaction is inhibited, apoptosis was activated.