RESUMO
Loss-of-function mutations in the ribonuclease angiogenin are associated with amyotrophic lateral sclerosis. Angiogenin has been shown to cleave transfer RNAs during stress to produce 'transfer-derived stress-induced RNAs'. Stress-induced tRNA cleavage is preserved from single-celled organisms to humans indicating it represents part of a highly conserved stress response. However, to date, the role of tRNA cleavage in amyotrophic lateral sclerosis remains to be fully elucidated. To this end, we performed small RNA sequencing on a human astrocytoma cell line to identify the complete repertoire of tRNA fragments generated by angiogenin. We found that only a specific subset of tRNAs is cleaved by angiogenin and identified 5'ValCAC transfer-derived stress-induced RNA to be secreted from neural cells. 5'ValCAC was quantified in spinal cord and serum from SOD1G93A amyotrophic lateral sclerosis mouse models where we found it to be significantly elevated at symptom onset correlating with increased angiogenin expression, imbalanced protein translation initiation factors and slower disease progression. In amyotrophic lateral sclerosis patient serum samples, we found 5'ValCAC to be significantly higher in patients with slow disease progression, and interestingly, we find 5'ValCAC to hold prognostic value for amyotrophic lateral sclerosis patients. Here, we report that angiogenin cleaves a specific subset of tRNAs and provide evidence for 5'ValCAC as a prognostic biomarker in amyotrophic lateral sclerosis. We propose that increased serum 5'ValCAC levels indicate an enhanced angiogenin-mediated stress response within motor neurons that correlates with increased survival. These data suggest that the previously reported beneficial effects of angiogenin in SOD1G93A mice may result from elevated levels of 5'ValCAC transfer RNA fragment.
RESUMO
Amyotrophic lateral sclerosis (ALS) presents a poorly understood pathogenesis. Evidence from patients and mutant SOD1 mouse models suggests vascular damage may precede or aggravate motor dysfunction in ALS. We have previously shown angiogenin (ANG) treatment enhances motor neuron survival, delays motor dysfunction and prevents vascular regression in the SOD1G93A ALS model. However, the existence of vascular defects at different stages of disease progression remains to be established in other ALS models. Here, we assessed vascular integrity in vivo throughout different disease stages, and investigated whether ANG treatment reverses vascular regression and prolongs motor neuron survival in the FUS (1-359) mouse model of ALS. Lumbar spinal cord tissue was collected from FUS (1-359) and non-transgenic control mice at postnatal day (P)50, P90 and P120. We found a significant decrease in vascular network density in lumbar spinal cords from FUS (1-359) mice by day 90, at which point motor neuron numbers were unaffected. ANG treatment did not affect survival or counter vascular regression. Endogenous Ang1 and Vegf expression were unchanged at P50 and P90; however, we found a significant decrease in miRNA 126 at P50, indicating vascular integrity in FUS mice may be compromised via an alternative pathway. Our study demonstrates that vascular regression occurs before motor neuron degeneration in FUS (1-359) mice, and highlights that heterogeneity in responses to novel ALS therapeutics can already be detected in preclinical mouse models of ALS.This article has an associated First Person interview with the joint first authors of the paper.