Familial dysautonomia model reveals Ikbkap deletion causes apoptosis of Pax3 ⁺ progenitors and peripheral neurons

Abstract

Familial dysautonomia (FD) is a devastating developmental and progressive peripheral neuropathy caused by a mutation in the gene inhibitor of kappa B kinase complex-associated protein (IKBKAP). To identify the cellular and molecular mechanisms that cause FD, we generated mice in which Ikbkap expression is ablated in the peripheral nervous system and identify the steps in peripheral nervous system development that are Ikbkap-dependent. We show that Ikbkap is not required for trunk neural crest migration or pathfinding, nor for the formation of dorsal root or sympathetic ganglia, or the adrenal medulla. Instead, Ikbkap is essential for the second wave of neurogenesis during which the majority of tropomyosin-related kinase A (TrkA + ) nociceptors and thermoreceptors arise. In its absence, approximately half the normal complement of TrkA + neurons are lost, which we show is partly due to p53-mediated premature differentiation and death of mitotically-active progenitors that express the paired-box gene Pax3 and give rise to the majority of TrkA + neurons. By the end of sensory development, the number of TrkC neurons is significantly increased, which may result from an increase in Runx3 + cells. Furthermore, our data demonstrate that TrkA + (but not TrkC + ) sensory and sympathetic neurons undergo exacerbated Caspase 3-mediated programmed cell death in the absence of Ikbkap and that this death is not due to a reduction in nerve growth factor synthesis. In summary, these data suggest that FD does not result from a failure in trunk neural crest migration, but rather from a critical function for Ikbkap in TrkA progenitors and TrkA + neurons.H ereditary sensory and autonomic neuropathies (HSANs) are a group of five phenotypically diverse but overlapping disorders of the peripheral nervous system (PNS) that result from mutations in 12 distinct genes (1). HSAN type 3, or familial dysautonomia (FD) (also called Riley-Day syndrome), results from an intronic mutation (IVS20 + 6T > C; 99.5% of patients) in a gene called inhibitor of kappa B kinase complex-associated protein or IKBKAP, causing mis-splicing and subsequent tissuespecific reductions in IKAP protein (2, 3). FD is marked by tachycardia, blood pressure lability, autonomic vomiting "crises," decreased pain and temperature sensation, and commonly death during early adulthood (4). The function of IKAP in the nervous system is unclear, nor is it understood why deletions in this broadly expressed gene primarily devastate the PNS. The earliest pathology study, performed on a 2-y-old child with FD, showed that ∼90% of cells in the dorsal root and sympathetic ganglia (SG) were missing (5). To identify IKAP's function in the developing PNS, we first need to establish the steps in which it is essential.The vertebrate PNS derives primarily from the neural crest, a multipotent, heterogeneous cell population that delaminates from the neural tube and migrates throughout the embryo (6). Those neural crest cells that stop laterally to the neural tube give rise to the chain of sensory...

A neuron autonomous role for the familial dysautonomia geneELP1in sympathetic and sensory target tissue innervation

“…Additional studies will be required to better understand whether Elp1 has any role in regulating filamin A function in Wnt1-specified mesenchymal cells during craniofacial bone and palate development. Although it seems possible that the developmental time points analyzed were not identical, our results differ from those obtained by George et al (George et al, 2013) who attributed Elp1-dependent sensory neuron loss at E12.5 to increased apoptosis caused by premature neuroblast cell cycle exit and differentiation. Although we could not confirm sensory or sympathetic neuron loss at E12.5, our results indicated that most neuron loss is likely due to target tissue innervation abnormalities in the absence of Elp1, rather than to relatively minor abnormalities in neurogenesis or differentiation.…”

“…Ablation of Elp1 in NCCs bypassed embryonic lethality that occurs in germline Elp1 knockout mice, and in agreement with previous results, it had no detectable role in their migration (George et al, 2013;Hunnicutt et al, 2012). In addition, we found that loss of Elp1 in NCCs did not alter sensory and sympathetic neuron proliferation or survival prior to their target tissue innervation, but at later stages of development profound abnormalities in neuron survival and target tissue innervation were identified.…”

“…Several studies have implicated Elp1 in cell migration, such as through its interaction with filamin A to regulate actin cytoskeleton reorganization during migration or by regulating α-tubulin acetylation in stabilized MTs, which are essential for neuron migration, neurite outgrowth and intracellular trafficking (Creppe et al, 2009;Solinger et al, 2010). However, previous studies in chick using shRNA to knockdown Elp1 in migrating NCCs (Hunnicutt et al, 2012) and more recent studies in mice that ablate Elp1 in migrating NCCs found no abnormalities in their migration (George et al, 2013). In agreement with these results, we found no changes in DRG or SCG location, neuron number, apoptosis or proliferation in the absence of Elp1 in E12.5 embryos after most NCC migration was complete.…”

“…However, many of the in vitro studies that have examined the functional consequences of Elp1 depletion have been performed in cellular contexts apart from that in which disease occurs, and in vivo studies to understand the function of Elp1 during mammalian sensory and sympathetic nervous system development have been hindered because loss of Elp1 in all tissues leads to embryonic lethality, which is not a common manifestation of the disease (Chen et al, 2009;Dietrich et al, 2011). Recent studies using shRNA to reduce Elp1 in chick NCCs (Hunnicutt et al, 2012) or to ablate it in mouse NCCs (George et al, 2013) indicate that Elp1 is not required for NCC migration, but how it functions in post-migratory neuron differentiation and/or target tissue innervation is still poorly understood.…”

Dimerization of elongator protein 1 is essential for Elongator complex assembly

“…Recently, Frances Lefcort and coworkers discovered that Elp1 is essential for the genesis of tropomyosinrelated kinase A nociceptors and thermoreceptors in mice (26). This role is consistent with the pain and temperature insensitivity phenotype of FD patients.…”