HAND2-mediated proteolysis negatively regulates the function of estrogen receptor α

A previous study demonstrated that the progesterone-inducible HAND2 gene product is a basic helix-loop-helix transcription factor and prevents mitogenic effects of estrogen receptor alpha (ER alpha) by inhibiting fibroblast growth factor signalling in mouse uteri. However, whether HAND2 directly affects the transcriptional activation function of ER alpha remains to be elucidated. In the present study, the physical interaction between HAND2 and ER alpha was investigating by performing an immunoprecipitation assay and an in vitro pull-down assay. The results demonstrated that HAND2 and ERa interacted in a ligand-independent manner. The in vitro pull-down assays revealed a direct interaction between HAND2 and the amino-terminus of ER alpha, termed the activation function-1 domain. To determine the physiological significance of this interaction, the role of HAND2 as a cofactor of ER alpha was investigated, which revealed that HAND2 inhibited the ligand-dependent transcriptional activation function of ER alpha. This result was further confirmed and the mRNA expression of vascular endothelial growth factor, an ER alpha-downstream factor, was decreased by the overexpression of HAND2. This inhibition of ligand-dependent transcriptional activation function of ER alpha was possibly attributed to the proteasomic degradation of ER alpha by HAND2. These results indicate a novel anti-tumorigenic function of HAND2 in regulating ER alpha-dependent gene expression. Considering that HAND2 is commonly hypermethylated and silenced in endometrial cancer, it is hypothesized that HAND2 may serve as a possible tumor suppressor, particularly in uterine tissue.