The Interaction between the Tyrosine Kinase Receptor EphA2 and RNF5: Structural Insights from an In Silico Approach

: EphA2 is a tyrosine kinase receptor from the Eph (erythropoietin-producing hepatocellular) family that is involved in several processes associated with regular cellular functions, as well as in different pathological conditions including cancer. EphA2 is up-regulated in many types of tumors, and the process of ligand-triggered receptor endocytosis and the subsequent degradation attract a lot of attention as a means of reducing receptor levels and its pro-oncogenic outcomes. In this context, the intracellular Sam (sterile alpha motif) domain of EphA2 (EphA2-Sam) plays an important role, being the region able to engage protein regulators of receptor endocytosis and stability. Recently, a novel cancer-related interaction involving the E3 ubiquitin ligase RNF5 and EphA2-Sam has been identified. Intriguingly, RNF5 is responsible for lowering receptor stability by targeting it for ubiquitination and consequent degradation. In certain types of breast cancer cells, down-regulation of EphA2 induced by RNF5 overexpression fosters pro-tumorigenic effects while RNF5 silencing and consequent EphA2 amplified regulation induces anticancer outcomes. Immunoprecipitation experiments point out that the membrane-binding domain of RNF5 interacts with EphA2-Sam, but detailed structural information on this protein-protein association is currently missing. To obtain fast insights into structural features governing the molecular recognition between RNF5 and EphA2, we set up an in silico approach combining structure predictions by the artificial intelligence tool AlphaFold and docking techniques. Such computational strategy, coupled with preliminary experimental validation with RNF5 peptide fragments, provides pilot information that can be exploited to develop original structure-based drug discovery approaches to identify novel anticancer compounds able to modulate the EphA2/RNF5 axis.