prior to this study , the affected index patient in family 1 was prescreened for smn1 mutations . whole exome sequencing ( wes ) was performed by axeq technologies ( seoul , south korea ) using the illumina ( san diego , ca ) trueseq kit to generate sequencing data . annotated wes data were examined for variants in genes selected for relevance to the phenotype . preceding sanger sequencing analysis of setx , als2 , sod1 , lmna , mfn2 , and trpv4 did not identify any pathogenic mutations in the affected individual of family 2 ; subsequently wes was carried out at the university of washington genome center . data from vcf files were analyzed using the in - house seave analysis pipeline ( kinghorn centre for clinical genomics , sydney , australia ) under homozygous and potentially compound heterozygote models . the effect of amino acid substitutions for sequence variants was assessed using the software sift and polyphen2 . sanger sequencing was performed to confirm the mutations in the probands and for segregation analysis in family members . two affected siblings were from a nonconsanguineous family of ashkenazi jewish origin ( figure 1 ) . in retrospect , she had difficulties performing sporting activities from the age of 15 years . she presented to medical attention at the age of 35 years with progressive lower limb weakness requiring the use of a walking stick . the impairment of motor function was progressive and at age 56 years she required a scooter and long arm crutch to mobilize . there were no significant ocular , bulbar , or sensory symptoms seen over the course of the illness and she had normal intellectual function . ( b ) schematic graph of the vrk1 coding region and the corresponding vrk1 protein shows the position of mutations identified in family 1 ( blue ) and 2 ( green ) . clinical examination was consistent with a profound symmetric distal muscle wasting affecting the upper and lower limbs symmetrically ( figure 2 ) . there was a predominantly distal pattern of weakness of greater severity in the lower than upper limbs , with relative preservation of proximal strength . neurophysiology showed preservation of sensory responses but severe and progressive reduction in compound motor action potential amplitudes with relative preservation of conduction velocities . motor evoked potentials from the upper limbs were prolonged with a central component suggesting upper motor neuron involvement in the disease , which is consistent with mri findings of atrophy of the spinal cord ( figure 3a ) . pontocerebellar hypoplasia or other developmental anomalies as reported in other patients previously described with vrk1 mutations were absent ( figure 3 ) . respiratory muscle weakness resulted in reduced mean inspiratory and expiratory pressures and a nonprogressive elevation of blood gas co2 . other respiratory parameters had been stable for 5 years with no requirement for assisted ventilation . serum creatine kinase was not elevated and an mri of the lower limbs showed profound proximal and distal muscle atrophy with fat replacement . there was relative preservation of the iliopsoas and adductor compartments bilaterally ( figure 3 ) . ( a ) mri of proband in family 1 ( sma ) . ( a.aa.c ) t2 images and ( a.d ) sagittal t1 image demonstrate mild nonspecific generalized atrophy and absence of pontocerebellar hypoplasia . ( a.e ) t2 image sagittal and ( a.f ) transverse spinal image demonstrate cord atrophy . ( a.ga.i ) t2 images of upper thigh , calf , and stir sequence of thigh demonstrate diffuse fatty replacement of muscle and relative sparing of adductor compartments . ( b.ab.c ) t2 images and ( b.d ) sagittal t1 image demonstrate absence of pontocerebellar hypoplasia . the proband has a similarly affected older brother with a slightly later onset of progressive motor weakness in the late teenage years , progression to requiring a walking stick in his early 30s , and the requirement for an articulated walking device and wheelchair in his late 40s . the patient was a 3-year - old girl with short stature and microcephaly who presented with initial distal lower limb muscle weakness and amyotrophy combined with pathologically brisk deep tendon reflexes and normal sensation . weakness progressed in a caudalocephalic direction , with loss of ambulation by 10 years of age and complete dependence for activities of daily living at 18 years . respiratory dysfunction developed with dyspnea and weak cough accompanied by reduction of forced vital capacity to below 40% predicted . neurophysiology studies at age 11 years demonstrated severe reduction in compound muscle action potential amplitudes combined with chronic and active denervation , consistent with motor neuropathy / neuronopathy . sensory responses were initially preserved but subsequently were reduced on repeat testing at age 19 years . the clinical possibility of juvenile als , with the presence of upper and lower motor neuron signs in more than 2 spinal regions and clinical progression , prompted sanger sequencing as described above . participating individuals were enrolled through the neurogenetics clinic concord hospital and the paediatric neuromuscular clinic at sydney children 's hospital randwick . these procedures were performed with informed consent according to protocols approved by the human ethics committees of sydney local health district , concord hospital ( hrec/11/crgh/105 ) , and south eastern sydney local health district ( hrec/13/powh/203 ) , australia . prior to this study , the affected index patient in family 1 was prescreened for smn1 mutations . whole exome sequencing ( wes ) was performed by axeq technologies ( seoul , south korea ) using the illumina ( san diego , ca ) trueseq kit to generate sequencing data . annotated wes data were examined for variants in genes selected for relevance to the phenotype . preceding sanger sequencing analysis of setx , als2 , sod1 , lmna , mfn2 , and trpv4 did not identify any pathogenic mutations in the affected individual of family 2 ; subsequently wes was carried out at the university of washington genome center . data from vcf files were analyzed using the in - house seave analysis pipeline ( kinghorn centre for clinical genomics , sydney , australia ) under homozygous and potentially compound heterozygote models . the effect of amino acid substitutions for sequence variants was assessed using the software sift and polyphen2 . sanger sequencing was performed to confirm the mutations in the probands and for segregation analysis in family members . two affected siblings were from a nonconsanguineous family of ashkenazi jewish origin ( figure 1 ) . in retrospect , she had difficulties performing sporting activities from the age of 15 years . she presented to medical attention at the age of 35 years with progressive lower limb weakness requiring the use of a walking stick . the impairment of motor function was progressive and at age 56 years she required a scooter and long arm crutch to mobilize . there were no significant ocular , bulbar , or sensory symptoms seen over the course of the illness and she had normal intellectual function . ( b ) schematic graph of the vrk1 coding region and the corresponding vrk1 protein shows the position of mutations identified in family 1 ( blue ) and 2 ( green ) . clinical examination was consistent with a profound symmetric distal muscle wasting affecting the upper and lower limbs symmetrically ( figure 2 ) . there was a predominantly distal pattern of weakness of greater severity in the lower than upper limbs , with relative preservation of proximal strength . neurophysiology showed preservation of sensory responses but severe and progressive reduction in compound motor action potential amplitudes with relative preservation of conduction velocities . motor evoked potentials from the upper limbs were prolonged with a central component suggesting upper motor neuron involvement in the disease , which is consistent with mri findings of atrophy of the spinal cord ( figure 3a ) . pontocerebellar hypoplasia or other developmental anomalies as reported in other patients previously described with vrk1 mutations were absent ( figure 3 ) . respiratory muscle weakness resulted in reduced mean inspiratory and expiratory pressures and a nonprogressive elevation of blood gas co2 . other respiratory parameters had been stable for 5 years with no requirement for assisted ventilation . serum creatine kinase was not elevated and an mri of the lower limbs showed profound proximal and distal muscle atrophy with fat replacement . there was relative preservation of the iliopsoas and adductor compartments bilaterally ( figure 3 ) . ( a.aa.c ) t2 images and ( a.d ) sagittal t1 image demonstrate mild nonspecific generalized atrophy and absence of pontocerebellar hypoplasia . ( a.e ) t2 image sagittal and ( a.f ) transverse spinal image demonstrate cord atrophy . ( a.ga.i ) t2 images of upper thigh , calf , and stir sequence of thigh demonstrate diffuse fatty replacement of muscle and relative sparing of adductor compartments . ( b.ab.c ) t2 images and ( b.d ) sagittal t1 image demonstrate absence of pontocerebellar hypoplasia . the proband has a similarly affected older brother with a slightly later onset of progressive motor weakness in the late teenage years , progression to requiring a walking stick in his early 30s , and the requirement for an articulated walking device and wheelchair in his late 40s . the patient was a 3-year - old girl with short stature and microcephaly who presented with initial distal lower limb muscle weakness and amyotrophy combined with pathologically brisk deep tendon reflexes and normal sensation . weakness progressed in a caudalocephalic direction , with loss of ambulation by 10 years of age and complete dependence for activities of daily living at 18 years . severe thoracolumbar scoliosis required surgical rod insertion at age 14 years . respiratory dysfunction developed with dyspnea and weak neurophysiology studies at age 11 years demonstrated severe reduction in compound muscle action potential amplitudes combined with chronic and active denervation , consistent with motor neuropathy / neuronopathy . sensory responses were initially preserved but subsequently were reduced on repeat testing at age 19 years . the clinical possibility of juvenile als , with the presence of upper and lower motor neuron signs in more than 2 spinal regions and clinical progression , prompted sanger sequencing as described above . participating individuals were enrolled through the neurogenetics clinic concord hospital and the paediatric neuromuscular clinic at sydney children 's hospital randwick . these procedures were performed with informed consent according to protocols approved by the human ethics committees of sydney local health district , concord hospital ( hrec/11/crgh/105 ) , and south eastern sydney local health district ( hrec/13/powh/203 ) , australia . testing identified compound heterozygous vrk1 mutations nm_003384.2 ( vrk1):c.(356a > g ) ; ( 1072c > t ) ( p.[h119r];[r358 * ] ) in the individuals with sma in family 1 and nm_003384.2 ( vrk1):c.(403 g > a ) ; ( 583t > g ) ( p.[g135r];[l195v ] ) in the patient with juvenile als in family 2 . no pathogenic variants were identified in other genes associated with sma or als in either patient or genes associated with short stature . the present cases with adult - onset distal sma and childhood motor neuron disease broaden the clinical spectrum of patients with vrk1 mutations . the identification of vrk1 mutations among these various motor phenotypes , with clinical overlap among juvenile als , dhmn + ps , and sma , may serve to further unite concepts of pathogenesis among motor neuron disorders . the results for family 1 are consistent with an autosomal recessive mode of inheritance with segregation of both vrk1 variants with the disease ( figure 1a ) . the c.1072c > t mutation is well - characterized and described as pathogenic in the ashkenazi jewish population when in a homozygous state . the c.356a > g variant ( esp frequency < 0.01% ) encodes a histidine residue that is conserved across multiple vertebrate species and is in the protein kinase domain of the vrk1 gene ( figure 1b ) . it was assessed with in silico tools ( polyphen2 score 1 and 0.999 , provean score 3.804 ) and is classified as damaging . the c.356a > g missense substitution was reported in one recent case of a 32-year - old man with a 5-year history of als without cerebellar hypoplasia . this sequence variant is most likely to be pathogenic as it is present in both the affected members of family 1 and a reported sporadic als case . in family 2 , both substituted amino acids are highly conserved across multiple species and the frequency of the novel c.583t > g mutation in public databases was 2 in 121382 . online database tools supported pathogenicity for c.403 g > a and c.583t > g mutations ( polyphen2 scores 1 and 0.997 , probably damaging ; sift score 0 and 0 , deleterious ; and cadd score 22.6 and 19.05 , respectively ) . the mutations were sanger sequenced for confirmation and each parent was found to carry one of the variants , supporting autosomal recessive inheritance ( figure 1 ) . three - dimensional structural protein analysis of the p.gly135arg vrk1 variant with the hope protein structure analysis suite database suggested a difference in charge , size , and hydrophobicity resulting in disturbances in local structure and protein misfolding . similarly , the p.leu195val vrk1 variant altered the amino acid size , causing an empty space in the core of the protein . gonzaga - jauregui et al . described childhood - onset distal sensory motor axonal neuropathy with microcephaly with a simplified gyral pattern and in a second case there was a similar motor and sensory axonal neuropathy with an underdeveloped cerebellar vermis . the latter case and this report suggest that vrk1 is not only important in development of motor system architecture , but are also important in the longer term in maintenance of motor neurons . vrk1 mutations should therefore be considered in the differential diagnosis of patients presenting with adult - onset sma and childhood motor neuron disease . marion stoll : organized and performed genetic testing on proband and family members ( family 1 ) , performed bioinformatics analysis and interpretation of wes data , writing of the manuscript . hooi ling teoh : collected data of the proband ( family 2 ) , writing of the manuscript . lee : clinically examined and interviewed the proband ( family 1 ) , writing of the manuscript . stephen reddel : clinically examined and performed neurophysiology on proband ( family 1 ) , critical revision of manuscript . ying zhu : supervised the creation of the bioinformatic pipeline for genomic analysis , assisted with bioinformatics analysis and interpretation of whole exome data , critical revision of manuscript . michael buckley : organized and performed genetic testing on proband and family members ( family 2 ) , interpretation of variant results , critical revision of manuscript . hugo sampaio : clinically examined and interviewed the proband ( family 2 ) , critical revision of manuscript . tony roscioli : obtained research funding for genomic sequencing , developed genomic consent forms for patient enrolment , supervised the creation of the bioinformatic pipeline for genomic analysis , performed bioinformatics analysis and interpretation of wes data , critical revision of manuscript . michelle farrar : provided clinical and scientific direction for the manuscript , drafted , reviewed and approved the final manuscript . garth nicholson : provided clinical and scientific direction for the manuscript , planned , drafted , reviewed , and finalized the manuscript , clinically examined , characterized , and interviewed the proband ( family 1 ) . supported by motor neurone disease research institute of australia . no industry , governmental , or institutional funding j. lee , s. reddel , y. zhu , m. buckley , h. sampaio , t. roscioli , m. farrar , and g. nicholson report no disclosures relevant to the manuscript .