Tumor necrosis factor receptor-associated factor 6 (TRAF6) deficiency results in exencephaly and is required for apoptosis within the developing CNS

Department of Pharmaceutical Sciences, Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada M5S 2S2.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 11/2000; 20(19):7384-93.
Source: PubMed


Tumor necrosis factor receptor-associated factors (TRAFs) are adaptor proteins important in mediating intracellular signaling. We report here that targeted deletion of traf6 greatly increases the frequency of failure of neural tube closure and exencephaly in traf6 (-/-) mice. The penetrance of this defect is influenced by genetic background. Neural tube fusion requires the coordination of several biological processes, including cell migration invoked by contact-dependent signaling, cell proliferation, and programmed cell death (PCD). To gain greater insight into the role of TRAF6 in these processes, neural development and migration within the CNS of traf6 (-/-) mice and controls were assessed through temporal examination of a number of immunohistochemical markers. In addition, relative levels of cellular proliferation and PCD were examined throughout embryonic development using bromodeoxyuridine (BrdU) and in situ terminal deoxynucleotidyl transferase-mediated dUTP biotinylated nick end labeling (TUNEL), respectively. The data suggest that loss of TRAF6 does not significantly alter the level of cellular proliferation or the pattern of neural differentiation per se, but rather regulates the level of PCD within specific regions of the developing CNS. Substantial reductions in TUNEL were observed within the ventral diencephalon and mesencephalon in exencephalic traf6 (-/-) embryos. Our results demonstrate a novel and prominent role for TRAF6 in the regional control of PCD within the developing CNS.

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Available from: Jeffrey Henderson, Jan 23, 2015
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    • "Notes contained in blue boxes relate to positive relationships, notes in red boxes relate to inverse or negative relationships. 1 Denny et al. (2013). 2 Girardi et al. (2008). 3 Lomaga et al. (2000); McGrath et al. (1999); McLin et al. (2008); Phelan et al. (2005); Regnier et al. (2002); Tissir et al. (2004). 4 Torchinsky et al. (1997); Hrubec et al. (2006). "
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    • "Almost all exencephaly and spina bifida is caused by failure of neural fold elevation, but the genetic causes of common human NTDs are not yet clarified owing to their genetic complexity (Greene and Copp, 2005; Greene et al., 2009; Harris and Juriloff, 1999, 2007; Juriloff and Harris, 2000). Although about 200 mutant mouse lines have been reported to cause NTDs, the etiologies of these defects are not fully understood (Greene and Copp, 2005; Greene et al., 2009; Harris and Juriloff, 2007; Lomaga et al., 2000). Therefore, studies of the mechanisms that lead to NTDs and the susceptibility genes for NTDs are meaningful not only biologically but also clinically. "
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    ABSTRACT: Histone methylation is the important transcription regulatory system that affects mammalian development and cell differentiation. Alterations in epigenetic gene regulation are associated with disease. Fbxl10 (F-box and leucine-rich repeat protein 10) is a JmjC domain-containing histone demethylase. Although Fbxl10 has been implicated in cell cycle regulation, cell death, senescence, and tumorigenesis, these functions are controversial and its physiological function is unclear. To determine the in vivo function of Fbxl10, in this study, we generated a homozygous mutation in the mouse Fbxl10 gene. About half of Fbxl10-deficient mice exhibit failure of neural tube closure, resulting in exencephaly and die shortly after birth. Fbxl10 deficiency also causes retinal coloboma and a curled tail with low penetrances. Fbxl10 mRNA is specifically expressed in the cranial neural folds at E8.5 embryos, and apoptosis increased in the neuroepithelium and mesenchyme of Fbxl10-deficient E9.5 embryos, consistent with neural tube defects found in Fbxl10-deficient mice. Depletion of Fbxl10 induced the increased expression of p19ARF, an inducer of apoptosis, in E8.5 embryos and mouse embryonic fibroblast cells. In addition, the number of mitotic neural progenitor cells is significantly increased in the mutant E14.5 brain. Our findings suggest that the Fbxl10 gene makes important contributions to embryonic neural development by regulating cell proliferation and cell death in mice.
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    • "In TRAF6 -/mice , an increase in the frequency of impaired neural tube closure and exencephaly has been found, suggesting that TRAF6 may regulate the levels of programmed cell death within specific regions of the developing CNS (Lomaga et al., 2000). In primary astrocytes stimulated with beta-amyloid, TRAF6 inhibition prevents reactive gliosis and iNOS expression (Akama & Van Eldik, 2000). "
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    ABSTRACT: The tumor necrosis factor (TNF) receptor-associated factors (TRAFs) were initially discovered as proteins that inducibly interact with the intracellular region of TNF receptors (TNFRs). Because the TNFRs lack intrinsic catalytic activity, the TRAFs are hypothesized to orchestrate signaling activation downstream of the TNFR superfamily, however their mechanism of activation remains unclear (Inoue et al., 2000; Bishop, 2004). Originally, the TRAFs were compared to the signal transducers and activators of transcription (STAT) protein family, due to their sequence homology, and the presence of multiple RING- and zinc-finger domains, suggesting that their function may be to regulate transcriptional activity (Rothe et al., 1994; Hu et al., 1994; Sato et al. 1995; Cheng et al., 1995). However, subsequent research focused predominantly on their cytoplasmic functions, and more recently on their function as E3 ubiquitin ligases (Pineda et al., 2007). In my research, I analyzed the subcellular localizations of the TRAFs following CD40 ligand (CD40L)-stimulation, and found that TRAF2 and 3 rapidly translocate into the nucleus of primary neurons and Neuro2a cells. Interestingly, similar analysis conducted in pre-B lymphocytes (Daudi cells) revealed a different response to CD40L-stimulation, with TRAF2 and 3 being rapidly degraded within 5-minutes of stimulation. These findings are significant because they demonstrate for the first time that the TRAFs translocate into the nucleus and suggest that they may function within the nucleus in a cell-specific manner. I next analyzed the ability of TRAF2 and 3 to bind to DNA, and found that they both bind to chromatin and the NF-kappaB consensus element in Neuro2a cells, following CD40L-stimulation. Similar analyses of the chromatin binding of TRAF2 and 3 in Daudi cells revealed that they were rapidly degraded, similar to the results from my analysis of their subcellular localization. These findings show for the first time that the TRAFs interact with DNA, and therefore support the hypothesis that the TRAFs may function within the nucleus as transcriptional regulators. Finally, I analyzed the ability of the TRAFs to regulate transcriptional activity by luciferase assay. Previous studies showed that overexpression of TRAF2 and 6 could induce NF-kappaB transcriptional activity; however researchers have not been able to determine the mechanism by which they do so. In my studies, I found that every TRAF can directly regulate transcriptional activity either as co-activators or co-repressors of transcription, in a cell- and target protein-specific manner. Additionally, I found that TRAF2 can act as a transcriptional activator, and that its ability to regulate transcription is largely dependent upon the presence of its RING-finger domain. In conclusion, these studies have revealed an entirely novel function for the TRAFs as immediate-early transcriptional regulators. Future research into the genes that are regulated by the specific TRAF complexes will further elucidate how the TRAFs regulate TNFR signaling, as well as whether dysfunctions in TRAF signaling may be associated with known disorders. If specific TRAF complexes are found to regulate specific genes, then pharmacological targeting of the individual TRAF complexes may allow for the highly specific inhibition of signaling events downstream of the TNFRs, without compromising overall receptor signaling, transcription factor pathways, or cellular systems.
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