Migration of sympathetic preganglionic neurons in the spinal cord is regulated by reelin‐dependent Dab1 tyrosine phosphorylation and CrkL

Abstract

The actions of Reelin in neuronal positioning in the developing cortex and cerebellum are relayed by Src-family kinase (SFK)-mediated phosphorylation of Dab1. Biochemical studies show that after phosphorylation Dab1 binds to an adaptor protein, CrkL. Whether CrkL is important for Reelin signaling in vivo is unknown, because crkl(-/-) embryos die before cortical development is complete. In the developing spinal cord, Reelin and components of its signaling pathway, VLDLR, ApoER2, and Dab1, control the positioning of sympathetic preganglionic neurons (SPN); however, it is not known whether SFKs or Dab1 tyrosine phosphorylation is required. In the present study, we asked whether Reelin-controlled SPN migration depends on tyrosine phosphorylation of Dab1 by SFKs and whether CrkL is involved in SPN migration. To answer these questions, we examined the location of SPN in various mutant mouse embryos. Results showed that, in dab1(5F/5F) embryos, which express a nonphosphorylated mutant of Dab1, and in src(-/-)fyn(-/-) double knockout embryos, the location of SPN is identical to that of reeler. These results show that tyrosine phosphorylation of Dab1 by SFKs is required for Reelin-regulated SPN positioning. In addition, we found that SPN migration in crkl(-/-) showed a partial reeler phenotype, suggesting a partial loss of response of SPN to Reelin signaling. These results suggest a role for CrkL in the Reelin signaling pathway to control neuronal migration.

Crk and Crk-Like Play Essential Overlapping Roles Downstream of Disabled-1 in the Reelin Pathway

“…CrkL -null embryos exhibit lethal defects in cardiac and neural crest development (Guris et al, 2001), precluding analysis of mature brain structures. However, there is a failure of preganglionic neuronal migration in the spinal cord mice lacking CrkL , but this defect is distinct from that seen in reeler (Yip et al, 2007). To analyze the tissue-specific contributions of Crk and CrkL to Reelin signaling, while avoiding the confounding effect of embryonic lethality, we generated floxed alleles for Crk and CrkL and crossed them with a Nestin-Cre transgene (Knoepfler et al, 2002) to obtain conditional mutations in Crk and CrkL in mouse neurons.…”

Reduction of Crk and CrkL expression blocks reelin-induced dendritogenesis

“…It remains to be determined whether Crk and CrkL are required for neuronal positioning in the neocortex downstream of Dab1 tyrosine phosphorylation. The CrkL mutant has some similarities to Reeler-Dab1 phenotypes in the spinal cord, but not in the neocortex (Guris et al, 2001;Yip et al, 2007). The Crk-null animals die around E16 with cardiac and craniofacial defects precluding examination for a Reeler-like phenotype (Park et al, 2006).…”

“…These molecules are known to operate on a number of signaling pathways, making assessment of their role in reelin signaling more complex. A CrkL mutant has recently been shown to have a partial similarity to the Reeler phenotype in the spinal cord, but it does not have Reeler-like phenotypes in other areas of the CNS (Guris et al, 2001;Yip et al, 2007). This might be due to compensation by highly similar adaptor molecules encoded by the Crk gene: CrkI and CrkII.…”

Reelin inhibits migration of sympathetic preganglionic neurons in the spinal cord of the chick

“…A recent study showed that Reelin induced phosphorylation of n-cofilin at the leading processes and stabilized the actin cytoskeleton of migrating cortical neurons (Chai et al, 2009). Reelin-dependent SPN migration involves the lipoprotein receptors VldlR and ApoER2 and Dab1 tyrosine phosphorylation by SFK and CrkL (Yip et al, 2004(Yip et al, , 2007. CrkL has been shown to bind C3G, an exchange factor for the small GTPase Rap1 (Ballif et al, 2004).…”

“…Tissues were cryoprotected with 30% phosphate-buffered sucrose and serially sectioned with a cryostat in the horizontal plane at a thickness of 20 lm. The morphology of migrating SPN in reelin electroporated embryos was studied using retrograde DiI labeling (Yip et al, 2007). reelin Electroporated embryos at S28 were fixed in 4% paraformaldehyde.…”