familial adult myoclonic epilepsy ( fame ) , or benign adult familial myoclonic epilepsy ( bafme ) , is a neurological disease of an autosomal - dominant inheritance , which is characterized clinically by adult onset of finger tremulous movement mixed with myoclonic jerks .
linkage studies have mapped fame - associated loci to chromosome 8q23.3-q24.11 in japanese families [ 1 , 2 ] , to chromosome 2p11.1-q12.2 in italian [ 35 ] and spanish   families , and chromosome 5p15.31-p15 in a french family .
although the identification of the causal gene(s ) for fame is of great importance to the understanding of the molecular basis of the disease , gene(s ) responsible for fame has not been identified . here ,
we report that a mutation of the gene encoding ubr5 ( ubiquitin protein ligase e3 component n - recognin 5 ) is associated with fame in a japanese family .
all of the family members were born in a small town in yamagata prefecture , about 400  km north of tokyo , japan .
the family consisted of 6 individuals with fame through four generations . both men and women
were affected with the disease , which is consistent with an autosomal - dominant inheritance .  
the proband ( iii-1 in figure 1 ) was a 49-year - old woman who had noticed a shivering - like involuntary movement of the bilateral hands since the age of around 25 years .
the shivering - like movement consisted of arrhythmic , fine twitches at the hands , which was exaggerated by posture , feeling of strain , and fatigue .
the involuntary movement was limited only in her hands , although it became slightly worsened with age , as compared with the condition at the onset .
no additional neurological symptoms , including cerebellar ataxia , dementia , muscle weakness , sensory disturbance , or other involuntary movements , were observed .
the other affected members of the family showed virtually identical symptoms to the proband , although her cousin ( 44-year - old woman : iii-4 in figure 1 ) had an episode of generalized seizure at the age of 42 years , and her 74-year - old father ( ii-4 ) and 72-year - old aunt ( ii-5 ) showed a progression of myoclonic involuntary movements from the hands to all four extremities with aging , resulting in some difficulties in walking with assistance .  
the study was approved by the medical ethics committee of yamagata university faculty of medicine . for exosome analysis , genomic dna from the proband
was extracted and purified from whole blood using qiaamp dna spin columns ( qiagen n.v .
exome capture was performed using the sureselect human all exon system ( agilent technologies , usa ) .
the manufacturer 's protocol for this system ( illumina paired - end sequencing library prep , version 1.0.1 ) was used , with the following modifications : 5  g of dna sample was fragmented by the nebnext dsdna fragmentase ( new england biolabs , usa ) .
sequencing reaction was performed on the illumina gaiix platform with version 4 chemistry and version 4 flow cells according to the manufacturer 's instructions .
the sequence reads to a reference human genome ( ucsc ncbi37/hg19 ) , we adopted the elandv2 software ( illumina inc .
snvs and indels detection were performed with run.pl script from casava v1.6 ( illumina inc .
snvs and indels were compared with ncbi dbsnp v131 to distinguish known snps ( those that had been deposited to dbsnp ) and novel snps ( those that were not in dbsnp ) .
all the snps were annotated by comparing their position to other genomic features including gene regions .
of 266,122 variants from ngs , 745 were expected to be functional ( i.e. , missense or nonsense ) . of these functional variants ,
finally , of 467 , we successfully designed 383 probes for goldengate assay ( illumina inc . ,
usa ) using the illumina assay design tool ( illumina design scores > 0.4 and designable ) . using this custom beadchip
, we searched disease - specific mutation among ten members of the present family with fame including five affected , 85 unrelated individuals from the same community , which is an ancestral population of the proband , and 24 individuals from a diverse ethnic panel commercially provided by the coriell cell repositories ( coriell institute for medical research , camden , nj , usa ) .
genomic dna from peripheral blood of the proband ( iii-1 ) was subjected to an exome analysis on ngs using the sureselect human all exon system .
all exons ( 37,354,942  bp ) of the proband genome were sequenced and 36,768,760  bp ( 98.43% ) were read . among them , nucleotide variations were observed at 266,122 sites containing 467 functional variants ( missense or nonsense ) .
of the 467 functional variants , the probes of 383 variants were successfully designed , and the goldengate assay was performed for 10 family members , consisting of 5 patients with fame ( ii-4 , ii-5 , iii-1 , iii-4 , and iv-2 ) and 5 nonaffected members ( ii-6 , iii-2 , iii-3 , iii-5 , and iv-1 ) ( figure 1 ) . of the 383 functional variants
examined , only c.5720g > a mutation ( p.arg1907his ) ( nm_015902.5 : exon 19 ) ( chr8:103293724:grch37/hg19 ) in the ubr5 gene ( gene i d : 51366 ) was found in all of the affected individuals in the family , but not in any of the non - affected family members ( figure 2 ) .
such mutation was not found in any of the unrelated 85 healthy japanese residents in the same community nor in any of the 24 individuals of various ethnicities .
the arg-1907 residue and its surrounding regions in ubr5 were highly conserved across species from homo sapiens to danio rerio ( figure 3 ) .
in the present study , an exome analysis of the proband and the subsequent goldengate assay for the family members identified the c.5720g > a mutation ( p.arg1907his ) in the ubr5 gene only in the affected members in the fame family , but not in the non - affected family members ( figure 2 ) .
no such mutation was found in any of the 109 persons examined , including the unrelated 85 healthy residents in the same community and the 24 subjects of various ethnicities .
the conservation of the arg-1907 residue in ubr5 across species suggests a functional importance of the residue ( figure 3 ) .
the ubr5 gene was reported to be located in chromosome 8q22.3 , which is close to the reported locus linked to japanese fame families . in european families with fame ,
chromosome 2p11.1-q12.2 [ 36 ] or chromosome 5p15.31-p15   has been mapped as the loci linked to fame .
therefore , three , or more , genes are assumed to be responsible for fame .
this genetic heterogeneity in fame is not surprising because in many genetic diseases including familial alzheimer 's disease and familial parkinsonism , a similar clinical phenotype can be produced by mutations in the different genes .
it seems likely that the proteins encoded by the causal genes for fame may functionally be related with each other , and each may converge to the same biochemical pathway .
the dysfunction of any of the fame - associated proteins may cause a dysfunction of the pathway , resulting in a similar clinical manifestation . 

ubr5 is a human homolog of the drosophila melanogaster tumor suppressor gene hyperplastic discs ( hyd ) [ 8 , 10 ] .
human ubr5 is a huge protein with 2799 aminoacid residues ( figure 2(a ) ) .
the protein is localized mainly in the nucleus and has been shown to be ubiquitously expressed in various human tissues , including the brain .
ubr5 has the hect ( homology to e6ap carboxy terminus ) domain at the c - terminus , which functions as e3 ubiquitin - protein ligase ( figure 2(a ) ) [ 811 ] .
e3 ubiquitin - protein ligases are involved in protein degradation in the ubiquitin - proteasome system , which plays an important role in a variety of fundamental cell regulations , such as gene expression , stress response , dna repair , and cell cycle .
previous studies have shown that mutations in the genes encoding e3 ubiquitin - protein ligases can cause hereditary neurological diseases including angelman syndrome   and autosomal recessive juvenile parkinsonism ( arjp ) , both of which show tremor or tremulous involuntary movement as in fame .   in conclusion
, we identified an fame - associated mutation in the ubr5 gene in the candidate region linked to japanese fame families .
further study is needed to clarify the significance of the mutant protein in the pathogenesis of fame .
 the causal gene(s ) for familial adult myoclonic epilepsy ( fame ) remains undetermined . to identify it , 
 an exome analysis was performed for the proband in a japanese fame family . 
 of the 383 missense / nonsense variants examined , only c.5720g > a mutation ( p.arg1907his ) in the ubr5 gene was found in all of the affected individuals in the family , but not in the nonaffected members . 
 such mutation was not found in any of the 85 healthy individuals in the same community nor in any of the 24 individuals of various ethnicities . 
 the present study demonstrated an fame - associated mutation in the ubr5 gene , which is located close to the reported locus linked to japanese fame families . 