This study provides four lines of evidence implicating the RNA polymerase II component ELP3 as critically important to the axonal biology of neurons and supporting the initial observation of involvement in human motor neuron disease. First, in an association study of 1483 individuals, ELP3 was associated with human motor neuron degeneration in the form of ALS in three different populations. Secondly, an independent mutagenesis screen in Drosophila for defects in neuronal communication and survival identified two different loss of function ELP3 mutations that each conferred abnormal photoreceptor axonal targeting and synaptic development, possibly signifying neurodegeneration. Thirdly, knockdown of ELP3 in zebrafish using antisense morpholino technology resulted in a dose-dependent shortening and abnormal branching of motor neurons with no concomitant morphological abnormality. Finally, risk-associated ELP3 alleles were associated with lower brain ELP3 expression in humans. These findings strongly implicate ELP3 in axonal biology and as a gene conferring risk of neuronal degeneration.
The published SNP-based genome-wide association studies in ALS have not detected associated ELP3
), but the protection-associated ELP3
microsatellite variants have a total frequency of ~11%, so approaches using tag-SNPs might not detect them. Although we too did not see any single SNP associations using a dense set of SNPs in and around ELP3
, we did identify a haplotype between microsatellite allele 6 and the C allele of rs12682496, suggesting either that an untyped causal variant lies on this haplotype or the haplotype itself is functional. Although in general microsatellites are not thought to be functional, differences in gene expression may be conferred by polymorphic microsatellites in regulatory regions (17
) or in coding sequences (20
). Consistent with an important genomic function, the D8S1820 microsatellite repeat is conserved within ELP3
in chimpanzees and rhesus monkeys, which suggests that it predates the common ancestor of apes and rhesus monkeys and is therefore at least 25 million years old.
The ELP3 protein is part of the RNA polymerase II complex and is involved in RNA processing (Supplementary Material, Table S5
). It contains an Fe4
cluster and is involved in histone acetylation (22
), RNA elongation (21
), modification of tRNA wobble nucleosides (23
) and an unknown catalytic function related to free radical reactions. Alteration in RNA processing is an element in the pathophysiology of several motor neuron disorders and neurodegenerative diseases, including ALS (2
), hereditary motor neuronopathies 5 [MIM600794] and 6 [MIM604320], Charcot–Marie–Tooth disease type 2D [MIM601472], spinal muscular atrophy [MIM253300], familial dysautonomia [MIM223900] (26
) and spinocerebellar ataxia 7 [MIM164500] (27
) (Supplementary Material, Table S5
). In addition, trinucleotide repeat neurodegenerative diseases have been proposed to be the result of a disruption of nuclear organization that prevents proper RNA processing.
A possible explanation for ELP3 involvement in motor neuron degeneration comes from its effect on transcription through histone acetylation. Heat shock proteins (HSPs) are molecular chaperones whose expression is increased in response to cellular stress. Motor neurons have a high threshold for activating HSPs, making them particularly vulnerable to stressors, including mutant SOD1
). ELP3 directly regulates HSP70 expression by acetylation of histones H3 and H4 (29
), and therefore one possible explanation for the association of high-expressing ELP3
alleles with protection from motor neuron degeneration in humans is the ability to increase the transcription of HSP70. Indeed, intraperitoneal injection of HSP70 prolonged the lifespan of G93A SOD1
transgenic mice (30
The association of ELP3 variants with motor neuron degeneration and axonal biology in general increases the evidence that the RNA processing pathway is of particular importance to neurons, and provides a potential therapeutic target for treatment of ALS.