The Eek receptor, a member of the Eph family of tyrosine protein kinases, can be activated by three different Eph family ligands

Abstract

The Eph family of receptors, the largest subgroup within the tyrosine protein kinase receptor family, are comprised of at least thirteen members, many of which are predominantly expressed in the developing and adult nervous system. In this study, we have isolated a fulllength cDNA, encoding the mouse homologue of a previous partially characterized Eek protein, a member of Eph receptor tyrosine kinase family. In a comparison of the amino acid sequences of various Eph family members, Eek is most similar to Ehk-3/MDK1, Sek/Cek8, Ehk-2, Hek/ Mek4/Cek4, and Bsk/Ehk1/Rek7/Cek7, which are predominantly expressed in the nervous system. Additionally, we have used a low-stringency PCR cloning technique to identify ligands, related to B61, that may interact with Eek. Three di erent GPI-linked ligands, namely Elf-1/ Cek7-L, Ehk1-L/E¯-2/Lerk3 and AL-1/RAGS, were isolated from mouse brain. To study the functional interactions between these ligands and the Eek receptors, we have constructed chimeric ligands consisting of the Fc portion of human IgG fused to their carboxyl-terminus. These chimeric ligands bound to, and activated both the Eek receptors and the Eek-TrkB chimeric receptors expressed in NIH3T3 cells. These ®ndings suggest that Eek receptor can be activated by at least three di erent GPI-linked ligands.

The EphA8 Receptor Regulates Integrin Activity through p110γ Phosphatidylinositol-3 Kinase in a Tyrosine Kinase Activity-Independent Manner

“…1A, bottom), it was evident that both EphA8-TrkB and EphA8 proteins were highly tyrosine phosphorylated due to the endogenous expression of EphA subgroup ligands as described previously (Fig. 1A, bottom, lanes 3 to 5) (30). In contrast, the EphA8 mutant containing Met in place of Lys-666, the putative ATP binding residue, was not tyrosine phos-phorylated (Fig.…”

“…To further exclude the possibility that EphA8 promotes integrin activity by activating ligand signaling pathways, NIH 3T3 cells were treated with PI-PLC, which eliminates GPI-linked ephrin A subgroup ligands from the cell surface as previously reported (30). We then compared the ability of the ephrin A-stripped cells to adhere to surfaces coated with fibronectin with or without preclustered ephrin A5-Fc soluble ligand stimulation (see Fig.…”

“…Binding assays using EphA5-Fc proteins and glutathione S-transferase (GST)-mixing experiments were performed as described previously (7,30). FarWestern blot assays were performed as described previously (11).…”

The p110γ PI‐3 kinase is required for the mechanism by which the EphA8‐induced neurites are modulated by ephrin‐A5 engagement

“…Prevous studies demonstrated that ephrinA5-Fc binds to the EphA8 receptor with high nM affinity (Park and Sanchez, 1997), and that treatment with its aggregated forms stimulates cell migration on the fibronectin substrate (Gu and Park, 2003). These studies clearly indicate that ephrin-A5 is an authentic ligand for EphA8.…”

Genetic analysis of the role of Eph receptors in the development of the mammalian nervous system

“…EphA8 receptors can bind at least three different type A ephrins (Park and Sµnchez 1997), but the one (or those) that play(s) a role in the formation of the projection from the superior colliculus to the contralateral inferior colliculus is (are) unclear. Engrailed homeobox genes regulate the expression of molecular cues (possibly ephrins) that direct growing retinal axons in the tectum to their correct topographic location.…”