From Dimerization to Signaling: Molecular Dynamics Insights into EphA1 and EphA2 Isoform Specificity

Abstract Ephrin receptors (Ephs) are receptor tyrosine kinases that regulate cellular growth, differentiation, and motility. EphA2, often overexpressed in cancer, is notable for its ligand-independent activation, which drives pro-oncogenic signaling distinct from the canonical, ligand-dependent pathway that restricts cell movement. While ligand binding induces extracellular clustering, kinase activation depends on dimerization within the transmembrane (TM) region. EphA1 and EphA2 differ substantially in their function, likely due to differences in their TM, juxtamembrane (JM), and membrane-proximal fibronectin type III (FN1/FN2) domains, but how these regions modulate dimerization remains unresolved. To address this, we performed extensive coarse-grained simulations using Martini 3 in an anionic POPC/PS/PIP2 membrane. Both receptors formed stable TM dimers, though EphA1 favored a symmetric AXXXGXXXG-centered interface, whereas EphA2 favoured an additional leucine zipper interface and both receptors sampled multiple configurations, reflecting substantial intrinsic variability. Basic residues in the JM region remained membrane-bound, and the EphA2 FN domain displayed sustained PIP2 interactions, consistent with previous observations. Notably, the FN2 domain alone restricted TM association in both receptors, whereas inclusion of the second FN1 domain restored dimerization but produced receptor-specific extracellular interfaces. These differences arise from distinct FN1-FN2 linker flexibilities and FN-domain membrane contacts, which together shape TM geometry and lipid engagement. Overall, our results highlight how TM and TM-proximal elements cooperatively tune dimerization in Eph receptors, offering mechanistic insight into the divergent activation behaviors of EphA1 and EphA2 and providing testable models relevant to cancer biology and Eph-driven signaling. Competing Interest Statement The authors have declared no competing interest.