Ehd3, a regulator of vesicular trafficking, is silenced in gliomas and functions as a tumor suppressor by controlling cell cycle arrest and apoptosis.

EHD3 [Eps15 homology (EH) domain-containing protein 3] is a protein that resides in tubular and vesicular membrane structures and participates in endocytic recycling, although all its functions are unknown. Since Ehd3 is most abundantly expressed in brain tissues, we examined its role in brain cancer progression. Using immunohistochemistry, we report loss of EHD3 expression in gliomas, including low-grade astrocytomas, suggesting that this is an early event in gliomagenesis. EHD3 expression is also very low in most of glioma cell lines tested. In two cell lines, a bisulfite sequencing method identifies promoter hypermethylation as a mechanism of Ehd3 silencing, and its expression was restored by the demethylating agent 5-Azacytidine. Doxycycline-inducible restoration of EHD3 expression to glioma cells decreases their growth and invasiveness and induces cell cycle arrest and apoptosis. Furthermore, shRNA-mediated Ehd3 silencing increases cell growth. Using a xenograft model, we demonstrate Ehd3 growth inhibitory functions in glioma cells in vivo. We suggest that Ehd3 functions as a tumor suppressor gene and loss of its expression is a very common event in gliomas. This is the first study to highlight the importance of a member of the C-terminal EHD proteins in cancer and to link their functions to the cell cycle and apoptosis.