A missense, loss-of-function YARS1 variant in a patient with proximal-predominant motor neuropathy

Abstract

CASE PRESENTATION

Aminoacyl-tRNA synthetases (ARSs) are a group of ubiquitously expressed, essential enzymes that are responsible for charging transfer RNAs (tRNAs) with cognate amino acids in the cytoplasm and mitochondria, a process critical for establishing fidelity in protein synthesis. Variants in the 37 ARS genes (all of which are encoded in the nucleus) have previously been associated with a spectrum of genetic disorders (Meyer-Schuman and Antonellis 2017; Kuo and Antonellis 2020). Tyrosyl-tRNA synthetase (encoded by YARS1) is responsible for charging tyrosyl-tRNA with tyrosine in the cytoplasm via a two-step aminoacylation reaction. Importantly, two YARS1 subunits must form an asymmetric homodimeric complex with two tyrosyl tRNAs for the aminoacylation reaction to occur (Ward and Fersht 1988). Previously, homozygosity or compound heterozygosity for loss-of-function YARS1 variants has been associated with severe, early-onset, multisystem disease, with phenotypes including liver dysfunction, developmental delay, brain anomalies, and sensorineural hearing loss (Nowaczyk et al. 2017; Williams et al. 2019; Averdunk et al. 2021; Estève et al. 2021). Furthermore, heterozygosity for missense variants or a small, in-frame deletion in YARS1 have been associated with dominant intermediate Charcot–Marie–Tooth neuropathy type C (DI-CMTC; MIM #608323), which is characterized by progressive weakness and sensory loss in the distal upper and lower extremities, with intermediate motor nerve conduction velocities between 25 and 45 m/sec (Jordanova et al. 2003, 2006; Hyun et al. 2013).

Here, we present a 47-yr-old female proband of European ancestry. She first noted symptoms at 18 yr of age when she developed tremors, weakness, and cramping of her hands. At 22, neuromuscular examination revealed asymmetric weakness in the distal upper extremities and proximal lower extremities. Reflexes were absent at the triceps, but were otherwise normal. Serial examinations have demonstrated slow progression of weakness. At 47, her exam showed asymmetric upper extremity weakness affecting both proximal and distal muscles and proximal > distal lower extremity weakness (Fig. 1A–C). Hip flexion and extension measured the weakest at 2/5 bilaterally on manual muscle testing. Atrophy was prominent in her forearms and hands (Fig. 1A). Deep tendon reflexes were absent except at the ankles, where hyporeflexia was noted. Sensory exam was normal to touch, vibration, position, and pinprick. Additional findings included rotary nystagmus (present since 18 yr of age), kyphoscoliosis, and tongue fasciculations. Creatine kinase was mildly elevated (947–1692 units/L) at symptom onset, but was reduced to 300 units/L at last measurement (47 yr of age). Electrodiagnostic studies performed at 23 yr of age demonstrated signs of widespread denervation in the upper and lower extremities and increased amplitude of motor unit potentials, suggesting abnormalities in motor neurons of the spinal cord. The motor and sensory nerve conduction velocities were normal. A left deltoid biopsy obtained at 21 yr of age showed grouped angular fibers, consistent with a neurogenic process. Clinical findings are summarized in Table 1.

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Figure 1.

A loss-of-function YARS1 variant in a patient with proximal-prominent neuropathy. (A–C) Clinical findings noted on the patient's physical exam, including atrophy of the distal upper extremities and hand muscles (A) and finger extensor weakness demonstrated by finger drop (B,C). (D) Pedigree of the proband's (arrow) family. Genotypes at p.Asp308 YARS1 are indicated below each individual (??/?? indicates an unknown genotype). Filled objects indicate that the individual is affected, whereas unfilled objects indicate that the individual is unaffected. A slash through an individual indicates that they are deceased. (E) YARS1 protein sequence alignment, including six evolutionarily diverse species centered around the affected p.Asp308 residue (indicated in red). (F) Haploid yeast lacking endogenous TYS1 were transformed with a LEU2-bearing pRS315 vector to express wild type, p.Gly41Arg, p.Asp308Tyr, or p.Pro167Thr TYS1, or with a vector containing no TYS1 insert (“No Insert”). Resultant yeast cultures were plated undiluted or diluted (1:10, 1:100, or 1:1000) on solid media without leucine or uracil (−leu −ura) (see Supplemental Fig. 1A) or on solid media containing 5-FOA and grown at 30°C. Replicates A and B indicate two separate colonies from a single transformation. Image shown is representative of two independent transformations using two different plasmid DNA preparations (p.Asp308Tyr TYS1 total n = 8). (G) Haploid yeast lacking endogenous TYS1 were transformed with a LEU2-bearing pAG425-GPD vector to express wild-type TYS1, p.Asp308Tyr TYS1, wild-type YARS1, or p.Asp308Tyr YARS1, or with a vector containing no TYS1 insert (“No Insert”). Resultant yeast cultures were plated undiluted or diluted (1:10, 1:100, or 1:1000) on solid media without leucine or uracil (−leu −ura) (see Supplemental Fig. 1B) or on solid media containing 5-FOA and grown at 30°C. Replicates A and B indicate colonies from transformations using two different plasmid DNA preparations. Image shown is representative of two independent transformations using two different plasmid DNA preparations (p.Asp308Tyr YARS1 total n = 6).

Based on the observations above, the patient was thought to have symptoms consistent with spinal muscular atrophy type III; however, genetic testing of SMN1 was normal (further genetic testing information provided below). Family history was negative for similar disease, and the patient's mother had no symptoms of neuropathy; however, the patient's father developed a sensory peripheral neuropathy in adulthood without symptoms of weakness. He did not undergo electrodiagnostic testing. The patient's two daughters (ages 20 and 22 yr) were examined by the proband's neuromuscular physician and had normal strength and reflexes.

TECHNICAL ANALYSIS

Based on clinical presentation, genetic testing was performed using the Invitae Comprehensive Neuropathies Gene Panel (Invitae), which evaluates 111 genes associated with hereditary neuropathies for (1) single-nucleotide variants and deletions/insertions <15 nt in length by targeted next-generation sequencing of coding exons plus 10–20 bp of flanking adjacent intronic sequence, and (2) duplications and deletions at single-exon resolution (Invitae 03200, https://www.invitae.com/en/providers/test-catalog/test-03200). This analysis identified a missense variant in YARS1 ({"type":"entrez-nucleotide","attrs":{"text":"NM_003680.4","term_id":"1893862349","term_text":"NM_003680.4"}}NM_003680.4:c.922G > T [p.Asp308Tyr]) (see Table 2). Importantly, familial testing demonstrated that this variant was absent in the patient's mother and two daughters, who are not affected with the neuropathy phenotype present in the patient (Fig. 1D). The patient's father was not available for genetic testing. Given these findings, it is possible that the variant observed in the proband may have arisen de novo, but further haplotype analysis would be required to rule out paternal inheritance.

VARIANT INTERPRETATION

DISCUSSION

In this study, we identified a previously unreported YARS1 variant that affects an amino acid residue in the anticodon binding domain. This variant was observed in a patient with a neuropathy phenotype that is clinically distinct from previously described YARS1-related dominant intermediate CMT (Table 1). If this variant is confirmed as pathogenic, this report will have expanded the allelic and phenotypic spectrum of YARS1-related dominant disease. Notably, a similar range of dominant neuropathy phenotypes have been described in association with loss-of-function missense variants and small, in-frame deletions in four other cytoplasmic, homodimeric ARSs (AARS1, GARS1, HARS1, and WARS1) (Antonellis et al. 2003; Latour et al. 2010; McLaughlin et al. 2012; Vester et al. 2013; Safka Brozkova et al. 2015; Meyer-Schuman and Antonellis 2017; Tsai et al. 2017).

As noted, the patient reported here has several phenotypic features not previously seen in patients with YARS1-associated DI-CMTC (Table 1). This case is unique in that the patient presented with upper extremity weakness, whereas all prior patients presented with lower extremity involvement prior to developing upper extremity symptoms; however, such upper limb phenotypes have been reported in patients with pathogenic GARS1 variants (Antonellis et al. 2003). The pattern of muscle weakness also differs from that seen in other patients with YARS1 variants. The patient presented here has predominant proximal lower extremity weakness with minimal distal weakness of these extremities. Although some patients with DI-CMTC have been reported to have proximal lower extremity weakness, it was reported as less severe than the distal weakness (Thomas et al. 2016). The patient reported here also lacks foot deformities and sensory abnormalities seen in most patients (Thomas et al. 2016; Nam et al. 2022). In addition, median motor nerve conduction velocities (NCVs) were normal on three separate electrodiagnostic studies in the third decade of life, whereas most previously reported adult DI-CMTC patients presented with reduced NCVs. The patient reported here was also identified to have persistent elevation of creatine kinase (CK) early in her disease course, which has not been reported in association with DI-CMTC and is rarely associated with CMT disease (Rudnik-Schöneborn et al. 2016).

To our knowledge, the p.Asp308Tyr variant described in this study is the first potentially pathogenic YARS1 variant identified in the anticodon binding domain; in contrast, all previously identified YARS1 variants map to the catalytic domain (Supplemental Fig. 1D). Importantly, variants in the anticodon binding domain of other ARSs have been associated with a more severe, atypical dominant neuropathy, including an infantile-onset spinal muscular atrophy caused by GARS1 variants (James et al. 2006; Eskuri et al. 2012; Markovitz et al. 2020). It will be interesting to test whether, and by what mechanism, mutations in the tRNA binding domain lead to differential clinical presentations.

In addition to aminoacylation activity, YARS1 has noncanonical functions that may be relevant for YARS1-related disease. First, a Drosophila model of YARS1-related CMT showed that YARS1 localizes to the nucleus and complexes with the scaffolding protein TRIM28 and histone deacetylase HDAC1 to enact a transcriptional regulation program, which is altered in the context of neurons expressing CMT-associated YARS1 variants (Bervoets et al. 2019). Second, noncanonical cytokine activity by secreted YARS1 cleavage products plays a role in angiogenesis and platelet biogenesis (Wakasugi et al. 2002; Yang et al. 2002; Greenberg et al. 2008). Finally, noncanonical cytoplasmic mRNA binding has been reported for many cytoplasmic ARSs in yeast, often with downstream consequences on post-transcriptional gene regulation (Levi and Arava 2019; Levi et al. 2020; Garin et al. 2021). Therefore, it is possible that dysfunction in noncanonical activities—in concert with any effects on tRNA charging—could explain the clinical heterogeneity observed in YARS1-related neuropathy. Further studies will be needed to fully understand the molecular mechanisms underlying the range of YARS1-related phenotypes.

ADDITIONAL INFORMATION

Supplementary Material

Footnotes

REFERENCES