Ephrin Type-A Receptor

      Ephrin Type-A Receptors (EphAs) represent the largest family of receptor tyrosine kinases in the mammalian genome. They regulate a number of signaling pathways through a number of downstream effectors.^[1] EphAs, which are typically localized to the dendritic spines of neurons, bind ephrins bound to the membranes of glial cells.^[2] Fourteen Eph receptors and eight ephrin ligands are encoded in the human genome.^[3] Upon binding an ephrin Eph receptors cluster together, undergo autophosphorylation, and trigger downstream signaling cascades that mediate cytoskeletal rearrangements and changes in cell adhesion resulting in dendritic spine retraction.^[4] The pathway connecting Eph activation and dendritic growth cone collapse has been well established. Upon Eph activation, ERK activity is inhibited leading to activation of TSC2. TSC2 inhibits RHEB which activates mTOR, a critical regulator of protein synthesis. As can be seen in the image at the left, ephrin stimulation reduces protein synthesis in neurons and triggers dendritic growth cone collapse.^[1] This has been validated in mouse models in which EphA4-knockout mice develop dendritic spines in the hippocampus that are significantly longer than present in wild type mice.^[2] A notable feature of Eph receptors and ephrins is that their interaction triggers bidirectional signals between the engaged cells. Forward signals are mediated by the Eph receptors in dendritic spines, while ephrins are mediate the reverse signals in glial astrocytes.^[2]

Involvement in Tuberous Sclerosis

      Tuberous sclerosis complex (TSC) is an autosomal dominant disease characterized by the presence of benign tumors called hamartomas. These benign tumors can affect virtually every organ system in the body.^[1] TSC is caused by mutations in the TSC1 or TSC2 genes. The loss of TSC2 and subsequent expression of constitutively active Rheb reduces the growth cone responsiveness to ephrin signals, resulting in dendritic spine abnormalities.^[1] Such abnormalities are often associated with neuropsychiatric symptoms, including intellectual disability, psychological deficits, and epilepsy. Additionally, 25-50% of individuals with TSC develop Autism.^[5]

Eph-Ephrin Interaction

      The extracellular part of Eph receptors includes the N-terminal ephrin (Ligand)-binding domain (LBD), a cysteine-rich domain (CRD), and two fibronectin Type-III Repeats (FN3).^[4] EphA binds ephrins with its LBD. Most ephrins have a similar rigid structure which includes four loops, AB, CD, FG, & GH. The LBD of EphA4 is said to be a “structural chameleon,” able bind both A and B class ephrins. This explains why Ephrin Type-A receptors exhibit cross-class reactivity.^[3] The overall structure of the EphA4 LBD includes four important loops, the BC, DE, GH, & JK loops. EphA4 binds the GH loop of the ephrin ligand within a deep pocket created by the EphA4 DE and JK loops. It is these loops, DE and JK, which undergo the greatest conformational shifts when binding either EphrinA2 or EphrinB2. When binding EphrinA2, EphA4-Arg 162 forms a hydrogen bond with EphrinA2-Leu 138, while EphA4-Met 164 and EphA4-Leu 166 participate in hydrophobic interactions with EphrinA2-Leu 138 and EphrinA2-PHe 136. Although EphA4 binds EphrinB2 in the same binding pocket, the local interactions are significantly different. Most notably, the alpha helix present in the EphA4-EphrinA2 JK loop is disrupted in the EphA4-EphrinB2 structure. This is due to the steric clash that would occur between EphrinB2-Trp 122 and EphA4 Met 164. Instead, EphA4-Arg 162 and EphrinB2-Trp 122 form hydrophobic stacking interactions among other interactions which stabilize the receptor-ligand complex.^[3] A morph of the movements EphA4 undergoes to bind EphrinA2 and EphrinB2 can be seen here.

      Eph-Ephrin complexes form two unique heterotetrameric assemblies consisting of distinct EphA2-EphA2 interfaces. The first tetrameric form is generated by Eph-Eph interactions only within the LBD. The second tetrameric form involves complex interactions in the LBD and in the region near the CRD.^[6] These two heterotetramers generate a continuous Eph-ephrin assembly when combined (Alternative Coloring). The proximity of kinase domains in an eph-ephrin tetramer, favors transphosphorylation of tyrosines in the cytoplasmic domains. Phosphorylation promotes kinase activity by orienting the activation segment of the kinase domain in a way that favors subsrate binding and subsequent signaling. It is this signaling that leads to growth cone collapse in dendritic spines, among other effects.^[4]