Specific association of estrogen receptor beta with the cell cycle spindleassembly checkpoint protein, MAD2

Estrogen receptors (ERs) are ligand-activated transcription factors that re gulate gene expression and cell growth. Two ERs now have been identified: E R alpha and the more recently discovered ER beta, The physiological functio n of ER beta remains unclear, but evidence from vascular injury studies and from ER beta knockout mice suggests that ER beta may be involved in the re gulation of cellular proliferation. Here we show a direct and specific inte raction between ER beta and the cell cycle mitotic spindle assembly checkpo int protein, MAD2 (mitosis arrest-deficient 2). The ER beta-MAD2 interactio n was identified by screening of a yeast two-hybrid system vascular endothe lial cell library with ER beta and confirmed with glutathione S-transferase -fusion protein interaction studies. In contrast, ER alpha did not interact with MAD2 in either the two-hybrid system or in the protein-protein intera ction experiments. Amino acids 173-208 in the hinge region of ER beta were sufficient to mediate the interaction with MAD2 in the two-hybrid system an d in glutathione S-transferase-fusion protein studies. These data identify a link between ER beta and MADZ of potential importance to regulation of th e cell cycle and support a function of ER beta distinct from the establishe d role of ERs as transcription factors.