HuD (ELAVL4) gain-of-function impairs neuromuscular junctions and induces apoptosis in in vitro and in vivo models of amyotrophic lateral sclerosis.

Early defects at the neuromuscular junction (NMJ) are among the first hallmarks of the progressive neurodegenerative disease amyotrophic lateral sclerosis (ALS). According to the "dying back" hypothesis, disruption of the NMJ not only precedes, but is also a trigger for the subsequent degeneration of the motoneuron in both sporadic and familial ALS, including ALS caused by the severe FUS pathogenic variant P525L. However, the mechanisms linking genetic and environmental factors to NMJ defects remain elusive. By taking advantage of co-cultures of motoneurons and skeletal muscle derived from human induced pluripotent stem cells (iPSCs), we show that the neural RNA binding protein HuD (ELAVL4) may underlie NMJ defects and apoptosis in FUS-ALS. HuD overexpression in motoneurons phenocopies the severe FUSP525L mutation, while HuD knockdown in FUSP525L co-cultures produces phenotypic rescue. We validated these findings in vivo in a Drosophila FUS-ALS model. Neuronal-restricted overexpression of the HuD-related gene, elav, produces per se a motor phenotype, while neuronal-restricted elav knockdown significantly rescues motor dysfunction caused by FUS. Finally, we show that HuD levels increase upon oxidative stress in human motoneurons and in sporadic ALS patients with an oxidative stress signature. On these bases, we propose HuD as an important player downstream of FUS mutation in familial ALS, with potential implications for sporadic ALS related to oxidative stress.

Emerging Roles for the RNA-Binding Protein HuD (ELAVL4) in Nervous System Diseases.

The main goal of this review is to provide an updated overview of the involvement of the RNA-binding protein (RBP) HuD, encoded by the ELAVL4 gene, in nervous system development, maintenance, and function, and its emerging role in nervous system diseases. A particular focus is on recent studies reporting altered HuD levels, or activity, in disease models and patients. Substantial evidence suggests HuD involvement in Parkinson's disease (PD), Alzheimer's disease (AD), and amyotrophic lateral sclerosis (ALS). Interestingly, while possible disease-causing mutations in the ELAVL4 gene remain elusive, a common theme in these diseases seems to be the altered regulation of HuD at multiple steps, including post-transcriptional and post-translational levels. In turn, the changed activity of HuD can have profound implications for its target transcripts, which are overly stabilized in case of HuD gain of function (as proposed in PD and ALS) or reduced in case of decreased HuD binding (as suggested by some studies in AD). Moreover, the recent discovery that HuD is a component of pathological cytoplasmic inclusion in both familial and sporadic ALS patients might help uncover the common molecular mechanisms underlying such complex diseases. We believe that deepening our understanding of the involvement of HuD in neurodegeneration could help developing new diagnostic and therapeutic tools.

ALS-related FUS mutations alter axon growth in motoneurons and affect HuD/ELAVL4 and FMRP activity.

Mutations in the RNA-binding protein (RBP) FUS have been genetically associated with the motoneuron disease amyotrophic lateral sclerosis (ALS). Using both human induced pluripotent stem cells and mouse models, we found that FUS-ALS causative mutations affect the activity of two relevant RBPs with important roles in neuronal RNA metabolism: HuD/ELAVL4 and FMRP. Mechanistically, mutant FUS leads to upregulation of HuD protein levels through competition with FMRP for HuD mRNA 3'UTR binding. In turn, increased HuD levels overly stabilize the transcript levels of its targets, NRN1 and GAP43. As a consequence, mutant FUS motoneurons show increased axon branching and growth upon injury, which could be rescued by dampening NRN1 levels. Since similar phenotypes have been previously described in SOD1 and TDP-43 mutant models, increased axonal growth and branching might represent broad early events in the pathogenesis of ALS.