HuD (ELAVL4) gain-of-function impairs neuromuscular junctions and induces apoptosis in familial and sporadic amyotrophic lateral sclerosis models

Abstract

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. However, the pathogenic mechanisms linking genetic and environmental factors to NMJ defects remain elusive. Here we show that increased expression of the neural RNA binding protein HuD (ELAVL4) at the presynaptic level is sufficient to cause NMJ defects and trigger apoptosis in co-cultures of motoneurons and skeletal muscle derived from human induced pluripotent stem cells (iPSCs). These phenotypes are strikingly similar to those caused by the severe FUS P525L variant. Moreover, HuD knock-down in motoneurons reverts the adverse effects of the FUSP525L mutation in iPSC-derived cocultures. These findings were validated in vivo by taking advantage of a Drosophila model, where neuronal-restricted overexpression of the HuD-related gene, elav, exacerbates the effects of FUS expression and produces per se a motor phenotype. Notably, in this fly model, neuronal-restricted elav knockdown significantly rescues motor dysfunction caused by FUS. Finally, we show that HuD levels are increased upon oxidative stress in human motoneurons, and in sporadic ALS patients with a signature related to oxidative stress. On these bases, we propose HuD as a key protein involved in causing early pathogenic alterations and offers a potential new therapeutic target for early intervention aimed at the NMJ in ALS.