Beteiligung der Ras-ähnlichen GTPase Rap2 an morphologischen Veränderungen dendritischer Dornfortsätze nach Stimulation des EphA4-Rezeptors
Zusammenfassung der Projektergebnisse
We have identified a novel EphA receptor signaling pathway that couples EphA4 activity with the regulation of the small GTPases, Rap1 and Rap2. This signaling pathway depends on an interaction of the carboxy terminus of EphA4 with the PDZ domain of the Rap GTPase-activating protein, SPAR. We found that ephrin stimulation plays a key role in SPAR-dependent Rap1 inactivation and that this signaling is important for growth cone collapse and downregulating integrin-mediated adhesion. We also demonstrated that Rap1 and Rap2 have differential effects on neuronal morphology and signaling downstream of EphA4. These results provide novel insight into signaling mechanisms that may be critical for neural circuitry formation and maintenance. Rap1 is well-known to promote neuronal differentiation and neurite outgrowth and has been recently implicated in the depolarization-induced growth of cortical dendrites and activity-dependent dendritic spine remodeling . A role for Rap1 in growth cone spreading has not been previously reported, but is consistent with the known role of Rap1 in promoting integrin-mediated adhesion, actin polymerization and cell spreading. Our data suggest that Rap1 inactivation decreases integrin-mediated adhesion downstream of EphA4 in neuronal cells. We also recently reported that EphA4 regulates dendritic spine morphology by influencing integrin activity. That growth cones and dendritic spines use some of the same mechanisms for remodeling is not surprising, given the importance of cell adhesion and the actin cytoskeleton for the architecture of both structures. Thus, we propose a model where decreased integrin activity through SPAR-mediated Rap1 inactivation contributes to ephrin-A-dependent growth cone collapse and decreased dendritic spine length and density. Our results suggest that Rap2 inactivation does not have as critical a role as Rap1 in ephrin-A- dependent growth cone collapse and inhibition of integrin activity in neuronal cells. Furthermore, we found that constitutively active Rap2 decreases growth cone size and dendritic spine length and density, whereas constitutively active Rap1 increases growth cone size and spine length . The dissimilar morphological effects of Rap1 and Rap2 on growth cones and dendritic spines are surprising since these two GTPases are closely related (~60% amino acid identity), share a number of the same regulators, and have some analogous functions. However, one of the amino acids in the effector domain is different between Rap1 and Rap2 , and some effectors may selectively mediate Rap1 or Rap2 signals. Active Rap1 has been shown to influence the subcellular localization of Rac and Rho regulatory proteins, leading to localized cell spreading and the formation of cell protrusions, consistent with positive effects of Rap1 on growth cone and dendritic spine size. On the other hand, Rap2 but not Rap1 interacts with TNIK (Traf2- and Nckinteracting kinase), a kinase expressed in the brain. Binding of activated Rap2 enhances the ability of TNIK to decrease F-actin levels and inhibit cell spreading. Such effects are consistent with the decreased size of growth cones and dendritic spines that we have observed in neurons transfected with constitutively active Rap2. The novel EphA4-SPAR-Rap1 pathway that we have identified likely acts in concert with other Ras and Rho pathways that are also activated downstream of EphA receptors to sculpt neuronal morphology and control the establishment and remodeling of neuronal connections. Because SPAR is a ubiquitously expressed protein and part of a family of three related proteins, the Eph-SPAR signaling connection may operate widely to regulate adhesion and morphology of many cell types through a combination of the different activities of Rap1 and Rap2.
Projektbezogene Publikationen (Auswahl)
- Eph receptors inactivate R-Ras through different mechanisms to achieve cell repulsion. J Cell Sci. 2006 Apr 1;119(Pt 7):1244-54. Epub 2006 Mar 7
Dail M, Richter M, Godement P, Pasquale EB
- The EphA4 receptor regulates dendritic spine remodeling by affecting β1-integrin signaling pathways. J Cell Biol. 2007 Sep 24; 178(7):1295- 307. Epub 2007 Sep 17
Bourgin C, Murai KK, Richter M, Pasquale EB
- The EphA4 Receptor Regulates Neuronal Morphology Through SPAR-Mediated Inactivation of Rap GTPases. J Neurosci. 2007 Dec 19;27(51):14205-15
Richter M, Murai KK, Bourgin C, Pak DT, Pasquale EB
