Familial Dysautonomia (FD) is a fatal disease characterized by degeneration of the peripheral nervous system (1, 2). FD patients carry a mutation in the gene IKBKAP resulting in reduced levels of the corresponding protein, IKAP, which assembles a multi-subunit complex called Elongator (3, 4). Although multiple functions for Elongator have been described, recent studies indicate that its primary role may lie in the modification of tRNAs that specifically recognize AA ending codons, and in the translation of mRNA transcripts that are enriched in these codons (5, 6). The crucial role of IKAP and Elongator in early development is demonstrated by the E10.5 lethality of Ikbkap ablation in the mouse embryo (7, 8). To circumvent this lethality and to study the function of IKAP in the cells most severely depleted in FD, we generated mice in which Ikbkap is selectively deleted in the neural crest lineage (9). These conditional knockout (CKO) mice exhibit all of the primary hallmarks of the human disease including a reduced number of afferents in the dorsal root ganglia (9). Analysis of sensory neurogenesis in mutant embryos shows that both neuronal progenitor cells and post-mitotic neurons exhibit increased expression of the pro-apoptosis protein p53. Elevated p53 levels are associated with multiple neurodegenerative diseases including ALS, Parkinson's, Alzheimer's, and Huntington's (10-16). Post-translational modifications, including ubiquitination and acetylation, are a primary regulator of p53 activity (17-19). Since numerous studies (5, 20-22), as well as data presented here, demonstrate a function for Elongator in protein acetylation, a primary thesis of this proposal is that acetylatio of p53 is altered in the absence of IKAP. Through multiple approaches we will determine whether altered levels of p53 isoforms are present in CKO embryos, as well as determine whether therapeutics that increase acetylation can rescue neuron death in the absence of IKAP. A second focus of the proposed work is to test our hypothesis that Elongator functions in tRNA modification and the translation of codon-biased proteins in mammalian neurons and glia. To this end we will test DRG cultures from mutant embryos for resistance to ?-toxin, a ribonuclease that specifically attacks Elongator-modified tRNAs, as well as quantify the expression of codon-biased fluorescent probes. Finally, we will test our preliminary data indicating that Brca2, a protein required for the proper acetylation of histones, is decreased in CKO embryos due to saturation of the Brca2 gene with AA ending codons. Thus, the proposed aims will not only identify molecular pathways that could be therapeutically targeted to slow the progressive nature of FD, but also shed light on the cellular function of Elongator in neuronal homeostasis. In addition, these studies bring the first neuroscience and developmental biology research programs to MSU Billings. This will provide tremendous additional research opportunities for MSU Billings students in our undergraduate research course, as well as add 18 paid undergraduate internships over the course of the grant.
Familial Dysautonomia (FD), a recessive fatal disease that devastates the peripheral nervous system, is caused by mutation in the gene IKBKAP that encodes a scaffolding protein for the multi-subunit complex, Elongator. Using a novel mouse model, the goal of this study is to gain a better understanding of Elongator's molecular and cellular function and how abrogation of this function leads to FD pathophysiology.
| Lefcort, Frances; Mergy, Marc; Ohlen, Sarah B et al. (2017) Animal and cellular models of familial dysautonomia. Clin Auton Res 27:235-243 |
| George, Lynn; Dunkel, Haley; Hunnicutt, Barbara J et al. (2016) In vivo time-lapse imaging reveals extensive neural crest and endothelial cell interactions during neural crest migration and formation of the dorsal root and sympathetic ganglia. Dev Biol 413:70-85 |
