A ternary switch determines ERα LBD conformation

The transcription factor estrogen receptor α (ERα) is the primary driver of ER+ breast cancer progression and a target of multiple FDA-approved anticancer drugs. Ligand-dependent activity of ERα is determined by the conformation of helix-12 (H12) within the ligand binding domain (LBD), but how H12 transitions from an unliganded (apo) state to active (estrogen-bound) or inactive (SERM/SERD-bound) states remains unresolved. Here, we present the first crystal structure of an apo ERα LBD, revealing a third distinct H12 conformation that regulates receptor activity. Structural mass-spectrometry, small-angle X-ray scattering, functional analysis and molecular dynamics simulations reveal that the apo conformation of H12 is stable in the absence of ligand, but is destabilised by Y537S and D538G breast cancer mutations driving constitutive activation. We propose a model in which H12 functions as a ternary molecular switch to determine receptor activity. These findings provide critical insights into the ligand-dependent and -independent regulation of ERα and have significant implications for therapeutic intervention.