Expression and regulation of Xenopus CRMP-4 in the developing nervous system

Abstract

KEY WORDS: Xenopus, primary neurogenesis, neurogenin, CRMP, NotchThe collapsin response mediator proteins (CRMPs) (also known as TOAD (turned on after division), Ulip (unc-33 like protein) and DRP (dihydropyrimidinase family) are a conserved family of cytosolic phosphoproteins highly expressed in the nervous system (Wang and Strittmatter, 1996). Even though the CRMPs exhibit more than 60% amino acid identity to the amidohydrolase family, they do not possess enzymatic activity (Wang and Strittmatter, 1997).The first CRMP was identified as an intracellular mediator of semaphorin/collapsin growth cone collapse (Goshima et al., 1995). However, numerous studies have demonstrated that the activities of CRMPs are not restricted to this repulsive guidance cue and participate in a broad spectrum of additional activities, with function being dependant on the specific interaction with various protein partners (Arimura et al., 2004). CRMP-2 participates in LPA-induced growth cone collapse and regulates axonogenesis through the binding of tubulin heterodimers (Inagaki et al., 2001;Fukata et al., 2002). CRMP-2 also contributes to the establishment of neuronal polarity through the association with Numb and promoting Numb-mediated endocytosis of the neuronal cell adhesion molecule L1 (Nishimura et al., 2003).Int. J. Dev. Biol. 51: 339-343 (2007) CRMP proteins are also targets of a variety of protein kinases. CRMP-2 and CRMP-4 were identified as brain-specific substrates for glycogen synthase kinase 3 (GSK3) and during growth cone collapse, phosphorylation by Rho-associated kinase inhibits microtubule assembly and Numb-mediated endoyctosis Yoshimura et al., 2005;Cole et al., 2006). Recently, CRMP-2 was identified as a negative regulator of p53 and it has been suggested to play a role in the regulation of proliferation (Llanos et al., 2006;Tahimic et al., 2006). Moreover, the CRMPs may contribute to the pathogenesis of specific neurodegenerative disorders (Charrier et al., 2003).Presently, we describe the expression analysis of CRMP-4, during early Xenopus embryogenesis. CRMP-4 is expressed in the differentiating primary neurons and later expression is maintained throughout the central nervous system and in the eye. Correspondingly, CRMP-4 is positively regulated by X-ngnr-1 and negatively regulated by the Notch pathway. Results and DiscussionXenopus CRMP-4 was identified in a microarray screen aimed at identifying genes induced in dissociated ectodermal explants by the neuronal determination transcription factor, X-ngnr-1. Comparison of the predicted amino acid sequence revealed Xenopus CRMP-4 is 90% identical to the corresponding human and mouse sequences. Lower identity was observed between Xenopus CRMP-2 (75%) and other mammalian CRMP members (71-74%) (Fig. 1). The Cdk5 priming and GSK3 phosphorylation sites identified in the mammalian CRMP-2 are conserved (Uchida et al., 2005;Yoshimura et al., 2005).The expression of CRMP-4 during early Xenopus embryogenesis was investigated by RT-PCR analysis with RNA isolated from var...

Xenopus zinc finger transcription factor IA1 (Insm1) expression marks anteroventral noradrenergic neuron progenitors in Xenopus embryos

“…Similar to the zebrafish (Lukowski et al, 2006), IA1 is strongly expressed in the primary nervous system. By in situ hybridization, IA1 transcripts are first detected at stage 11-11.5 in precursors of the primary neurons and of the trigeminal placodes, after the bHLH gene Ngnr1 (Ma et al, 1996) already activated at stage 10.5 and before ElrC and terminal differentiation markers such as CRMP-4 (Carruthers et al, 2003;Souopgui et al, 2007) (Fig. 1A,B and data not shown).…”

Dihydropyrimidinase‐like protein 3 expression is negatively regulated by MYCN and associated with clinical outcome in neuroblastoma

“…Dihydropyrimidinase‐like proteins were first characterized as phosphoproteins that were highly expressed in the developing nervous system and DPYSL family members regulate neurite outgrowth, axonal guidance and cell differentiation . Recently, a study on the roles of DPYSLs in vivo indicated that during neurulation in zebrafish DPYSL3 was required for proper positioning of neural crest cells in the developing spinal cord .…”

“…Dihydropyrimidinase-like proteins were first characterized as phosphoproteins that were highly expressed in the developing nervous system and DPYSL family members regulate neurite outgrowth, axonal guidance and cell differentiation. (13,29,30) Recently, a study on the roles of DPYSLs in vivo indicated that during neurulation in zebrafish DPYSL3 was required for proper positioning of neural crest cells in the developing spinal cord. (31) This report highlights the importance of DPYSL3 in differentiation of neural crest cells and supports our findings that DPYSL3 expression may be important during the differentiation of neural crest-derived NB tumor cells.…”

Ascl1 as a Novel Player in the Ptf1a Transcriptional Network for GABAergic Cell Specification in the Retina

“…Digoxigenin-labeled antisens RNA probes for gad1, VGlut1 [56], TH [53], tubb2b [48], Bru [57], dpysl3 [58], Ptf1a [45] and Ascl1 [50] were generated according to the manufacturer’s instruction (Roche). Whole mount in situ hybridization [59] and double fluorescent in situ hybridizations on cryosections [60] were carried out as previously described.…”