The RNA-Binding Protein HuD Binds Acetylcholinesterase mRNA in Neurons and Regulates its Expression after Axotomy

Abstract

After axotomy, expression of acetylcholinesterase (AChE) is greatly reduced in the superior cervical ganglion (SCG); however, the molecular events involved in this response remain unknown. Here, we first examined AChE mRNA levels in the brain of transgenic mice that overexpress human HuD. Both in situ hybridization and reverse transcription-PCR demonstrated that AChE transcript levels were increased by more than twofold in the hippocampus of HuD transgenic mice. Additionally, direct interaction between the HuD transgene product and AChE mRNA was observed. Next, we examined the role of HuD in regulating AChE expression in intact and axotomized rat SCG neurons. After axotomy of the adult rat SCG neurons, AChE transcript levels decreased by 50 and 85% by the first and fourth day, respectively. In vitro mRNA decay assays indicated that the decrease in AChE mRNA levels resulted from changes in the stability of presynthesized transcripts. A combination of approaches performed using the region that directly encompasses an adenylate and uridylate (AU)-rich element within the AChE 3Ј-untranslated region demonstrated a decrease in RNA-protein complexes in response to axotomy of the SCG and, specifically, a decrease in HuD binding. After axotomy, HuD transcript and protein levels also decreased. Using a herpes simplex virus construct containing the human HuD sequence to infect SCG neurons in vivo, we found that AChE and GAP-43 mRNA levels were maintained in the SCG after axotomy. Together, the results of this study demonstrate that AChE expression in neurons of the rat SCG is regulated via post-transcriptional mechanisms that involve the AU-rich element and HuD.

Characterization of Multiple Exon 1 Variants in Mammalian HuD mRNA and Neuron-Specific Transcriptional Control via Neurogenin 2

“…Semiquantitative PCR and quantification was performed as previously described (Deschênes-Furry et al, 2007). Annealing temperature and PCR cycle numbers within linear range of amplification were first determined for each primer set (Table 1).…”

“…The developmental and localized expression of HuD in the brain is reflective of its established roles in neuronal differentiation and function (for review, see Deschênes-Furry et al, 2006). In addition, HuD has also been implicated in neuronal regeneration following axotomy (Anderson et al, 2003;Deschênes-Furry et al, 2007) or seizure Tiruchinapalli et al, 2008), learning and memory (Quattrone et al, 2001), and numerous pathologies, including Alzheimer's and Parkinson's disease (Noureddine et al, 2005;Amadio et al, 2009), and spinal muscular atrophy (Hubers et al, 2011). At the intracellular level, HuD typically binds to U-rich elements in the 3Ј untranslated region (UTR) and poly(A) tails to regulate transcript stability, transport, and translation.…”

“…Our study is also the first to identify a positive regulatory event that induces HuD mRNA expression. Given that expression of HuD is altered during learning and memory (Quattrone et al, 2001), nerve regeneration (Anderson et al, 2003;Deschênes-Furry et al, 2007), and neurological disorders (Noureddine et al, 2005;Bolognani et al, 2007;Tiruchinapalli et al, 2008;Amadio et al, 2009;Hubers et al, 2011), it seems reasonable to postulate that under these conditions, the expression and/or function of E1 variants are specifically affected. Moreover, Ngn2 has also been linked to learning and memory (Galichet et al, 2008) and neurological disorders (Uhrig et al, 2009), thereby raising the interesting possibility that Ngn2-dependent HuD transcription is causally or casually involved in these physiological and pathological conditions.…”

Emerging complexity of the HuD/ELAVl4 gene; implications for neuronal development, function, and dysfunction

“…Additionally, HuD has been implicated in axonal regeneration in two models of nerve injury. A study in our laboratory demonstrated that, between 1 and 4 d following axotomy of superior cervical ganglion (SCG) neurons, HuD protein and mRNA levels, its interaction with AChE mRNA, and AChE transcript levels decreased (Deschenes-Furry et al 2007). Importantly, the reduction in AChE mRNA abundance could be rescued by localized overexpression of HuD delivered via a viral vector.…”

Activity-dependent expression of RNA binding protein HuD and its association with mRNAs in neurons

“…These neuronal mRNAs involved in neuronal differentiation and synaptic plasticity contain AU-rich elements (AREs) in their 3' untranslated regions (3' UTR), [25][26][27][28][29][30][31] such as those encoding GAP-43, Neurosperin and Acetylcholinesterase mRNA. Previous studies demonstrated the importance of cytoplasmic localization of HuD RBP in differentiating PC12 cells.…”