Mutations in UBQLN2 cause dominant X-linked juvenile and adult-onset ALS and ALS/dementia

Journal name:
Date published:
Published online

Amyotrophic lateral sclerosis (ALS) is a paralytic and usually fatal disorder caused by motor-neuron degeneration in the brain and spinal cord. Most cases of ALS are sporadic but about 5–10% are familial. Mutations in superoxide dismutase 1 (SOD1)1, 2, TAR DNA-binding protein (TARDBP, also known as TDP43)3, 4 and fused in sarcoma (FUS, also known as translocated in liposarcoma (TLS))5, 6 account for approximately 30% of classic familial ALS. Mutations in several other genes have also been reported as rare causes of ALS or ALS-like syndromes7, 8, 9, 10, 11, 12, 13, 14, 15. The causes of the remaining cases of familial ALS and of the vast majority of sporadic ALS are unknown. Despite extensive studies of previously identified ALS-causing genes, the pathogenic mechanism underlying motor-neuron degeneration in ALS remains largely obscure. Dementia, usually of the frontotemporal lobar type, may occur in some ALS cases. It is unclear whether ALS and dementia share common aetiology and pathogenesis in ALS/dementia. Here we show that mutations in UBQLN2, which encodes the ubiquitin-like protein ubiquilin2, cause dominantly inherited, chromosome-X-linked ALS and ALS/dementia. We describe novel ubiquilin2 pathology in the spinal cords of ALS cases and in the brains of ALS/dementia cases with or without UBQLN2 mutations. Ubiquilin2 is a member of the ubiquilin family, which regulates the degradation of ubiquitinated proteins. Functional analysis showed that mutations in UBQLN2 lead to an impairment of protein degradation. Therefore, our findings link abnormalities in ubiquilin2 to defects in the protein degradation pathway, abnormal protein aggregation and neurodegeneration, indicating a common pathogenic mechanism that can be exploited for therapeutic intervention.

At a glance


  1. Mutations of UBQLN2 in patients with ALS and ALS/dementia.
    Figure 1: Mutations of UBQLN2 in patients with ALS and ALS/dementia.

    a, The mutation c.1490C>A, resulting in p.P497H, was identified in a large family with ALS (family 186). This family was used to map X-linked ALS. The pedigree is shown on the left and DNA sequences are shown on the right: wild-type sequence (upper panel) and a representative hemizygous mutation in a male patient, V3 (lower panel). All affected members whose DNA samples were available for sequencing had the mutation. Two obligate carriers (III4 and IV2) were identified as having the same mutation. For simplicity and clarity, more than one unaffected individuals of both genders are represented by a single diamond and more than one unaffected male individual is represented by a single square. Filled symbols, affected individuals; open symbols, unaffected individuals; m, individuals with a mutation in UBQLN2; n, individuals without a mutation in UBQLN2. b, The mutation c.1516C>A (p.P506T) was identified in family 6316: the pedigree is shown in the left panel and sequences in the right panel (showing a heterozygous mutation from a female obligate carrier, II1). In a and b, probands are indicated with arrows and patients with dementia are indicated with asterisks. c, Evolutionary conservation of amino acids in the mutated region of ubiquilin2 in various species. Comparison of human (Homo sapiens) ubiquilin 2 and its orthologues in chimpanzee (Pan troglodytes), dog (Canis lupus familiaris), cattle (Bos taurus), mouse (Mus musculus) and rat (Rattus norvegicus). Amino acids identical to those in the human protein are shown in black and non-identical ones are in red. The positions of the C-terminal amino acids are shown on the right. Mutated amino acids are indicated by arrows. d, Predicted structural and functional domains of ubiquilin2, a protein of 624amino acids. Predicted structural and functional domains include a ubiquitin-like domain (UBL, 33–103), four heat-shock-chaperonin-binding motifs (STI1), twelve PXX repeats (491–526) and a ubiquitin-associated domain (UBA). ALS- and ALS/dementia-linked mutations are clustered in the 12 PXX repeats.

  2. Ubiquilin-2-immunoreactive inclusions in the spinal cord and hippocampus.
    Figure 2: Ubiquilin-2-immunoreactive inclusions in the spinal cord and hippocampus.

    ag, Spinal cord (ac) and hippocampal (dg) sections from a patient with a UBQLN2P506T mutation were analysed with confocal microscopy (ac) and immunohistochemistry (dg), using a monoclonal antibody against ubiquilin2 (ubiquilin2-C). The ubiquilin-2-positive and skein-like inclusions (arrowhead) are shown in a spinal motor neuron (a). These inclusions are also ubiquitin-positive (b, c). In the hippocampus, the ubiquilin-2-positive inclusions are shown in the molecular layer of the fascia dentata (d, e), CA3 (f) and CA1 (g). White arrows in d indicate the middle region of the molecular layer with ubiquilin-2-positive inclusions. A higher-magnification image of the boxed area in d is shown in e. Black arrows indicate representative inclusions in neurites (eg), and arrowheads indicate cytoplasmic inclusions in the cell bodies (f and g). Scale bars: ac, 10μm; d, 200μm; e, 50μm; f and g, 25μm.

  3. Co-localization of ubiquilin[thinsp]2 with ALS- and dementia-linked TDP43.
    Figure 3: Co-localization of ubiquilin2 with ALS- and dementia-linked TDP43.

    al, Neuro-2a cells were transfected with various combinations of wild-type (WT) ubiquilin2, mutant (M) ubiquilin2 (P497H), wild-type TDP43 and a C-terminal fragment of TDP43 (amino acids 218–414) that is linked to ALS and FTLD. Ubiquilin2 is GFP-tagged and TDP43 is mCherry-tagged. Wild-type and mutant ubiquilin2 are mostly cytoplasmic. Wild-type TDP43 is located almost exclusively in the nuclei and C-TDP43 is almost exclusively cytoplasmic. TDP43 inclusions are co-localized with wild-type (gi) and mutant (P497H) (jl) ubiquilin2 (arrows). Some ubiquilin-2-positive inclusions are TDP43-negative (arrowhead). Scale bars, 10μm.

  4. Mutations in ubiquilin[thinsp]2 lead to ubiquitin-mediated impairment of proteasomal degradation.
    Figure 4: Mutations in ubiquilin2 lead to ubiquitin-mediated impairment of proteasomal degradation.

    a, b, UbG76V–GFP fluorescence intensity (arbitrary units, a.u.) was quantified by flow cytometry 48h after transient transfection of Neuro-2a cells with either wild-type (WT) or mutant (P497H or P506T) UBQLN2 (a). The dynamics of UbG76V–GFP reporter degradation after blockage of protein synthesis with cycloheximide for 0, 2, 4, and 6h are shown in b. Rates of UPS-reporter degradation were significantly slower for both the P497H and P506T mutants when compared to the wild-type at 4h and 6h. Mean fluorescence before cycloheximide administration was standardized as 100%. Data are averaged from at least three independent experiments. *, P<0.05; **, P<0.01; ***, P<0.001 (indicating significant differences when compared to wild-type by two-tailed Student’s t-test). Error bars, means±s.e.m.


  1. Deng, H. X. et al. Amyotrophic lateral sclerosis and structural defects in Cu,Zn superoxide dismutase. Science 261, 10471051 (1993)
  2. Rosen, D. R. et al. Mutations in Cu/Zn superoxide dismutase gene are associated with familial amyotrophic lateral sclerosis. Nature 362, 5962 (1993)
  3. Kabashi, E. et al. TARDBP mutations in individuals with sporadic and familial amyotrophic lateral sclerosis. Nature Genet. 40, 572574 (2008)
  4. Sreedharan, J. et al. TDP-43 mutations in familial and sporadic amyotrophic lateral sclerosis. Science 319, 16681672 (2008)
  5. Kwiatkowski, T. J., Jr et al. Mutations in the FUS/TLS gene on chromosome 16 cause familial amyotrophic lateral sclerosis. Science 323, 12051208 (2009)
  6. Vance, C. et al. Mutations in FUS, an RNA processing protein, cause familial amyotrophic lateral sclerosis type 6. Science 323, 12081211 (2009)
  7. Chen, Y. Z. et al. DNA/RNA helicase gene mutations in a form of juvenile amyotrophic lateral sclerosis (ALS4). Am. J. Hum. Genet. 74, 11281135 (2004)
  8. Greenway, M. J. et al. ANG mutations segregate with familial and ‘sporadic’ amyotrophic lateral sclerosis. Nature Genet. 38, 411413 (2006)
  9. Nishimura, A. L. et al. A mutation in the vesicle-trafficking protein VAPB causes late-onset spinal muscular atrophy and amyotrophic lateral sclerosis. Am. J. Hum. Genet. 75, 822831 (2004)
  10. Yang, Y. et al. The gene encoding alsin, a protein with three guanine-nucleotide exchange factor domains, is mutated in a form of recessive amyotrophic lateral sclerosis. Nature Genet. 29, 160165 (2001)
  11. Chow, C. Y. et al. Deleterious variants of FIG4, a phosphoinositide phosphatase, in patients with ALS. Am. J. Hum. Genet. 84, 8588 (2009)
  12. Maruyama, H. et al. Mutations of optineurin in amyotrophic lateral sclerosis. Nature 465, 223226 (2010)
  13. Ticozzi, N. et al. Paraoxonase gene mutations in amyotrophic lateral sclerosis. Ann. Neurol. 68, 102107 (2010)
  14. Mitchell, J. et al. Familial amyotrophic lateral sclerosis is associated with a mutation in D-amino acid oxidase. Proc. Natl Acad. Sci. USA 107, 75567561 (2010)
  15. Johnson, J. O. et al. Exome sequencing reveals VCP mutations as a cause of familial ALS. Neuron 68, 857864 (2010)
  16. Lansbury, P. T. & Lashuel, H. A. A century-old debate on protein aggregation and neurodegeneration enters the clinic. Nature 443, 774779 (2006)
  17. Deng, H. X. et al. FUS-immunoreactive inclusions are a common feature in sporadic and non-SOD1 familial amyotrophic lateral sclerosis. Ann. Neurol. 67, 739748 (2010)
  18. Neumann, M. et al. Ubiquitinated TDP-43 in frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Science 314, 130133 (2006)
  19. Shibata, N. et al. Intense superoxide dismutase-1 immunoreactivity in intracytoplasmic hyaline inclusions of familial amyotrophic lateral sclerosis with posterior column involvement. J. Neuropathol. Exp. Neurol. 55, 481490 (1996)
  20. Mackenzie, I. R. et al. Pathological TDP-43 distinguishes sporadic amyotrophic lateral sclerosis from amyotrophic lateral sclerosis with SOD1 mutations. Ann. Neurol. 61, 427434 (2007)
  21. Neumann, M. et al. Frontotemporal lobar degeneration with FUS pathology. Brain 132, 29222931 (2009)
  22. Urwin, H. et al. FUS pathology defines the majority of tau- and TDP-43-negative frontotemporal lobar degeneration. Acta Neuropathol. 120, 3341 (2010)
  23. Nonaka, T., Kametani, F., Arai, T., Akiyama, H. & Hasegawa, M. Truncation and pathogenic mutations facilitate the formation of intracellular aggregates of TDP-43. Hum. Mol. Genet. 18, 33533364 (2009)
  24. Ko, H. S., Uehara, T., Tsuruma, K. & Nomura, Y. Ubiquilin interacts with ubiquitylated proteins and proteasome through its ubiquitin-associated and ubiquitin-like domains. FEBS Lett. 566, 110114 (2004)
  25. Dantuma, N. P., Lindsten, K., Glas, R., Jellne, M. & Masucci, M. G. Short-lived green fluorescent proteins for quantifying ubiquitin/proteasome-dependent proteolysis in living cells. Nature Biotechnol. 18, 538543 (2000)
  26. Kay, B. K., Williamson, M. P. & Sudol, M. The importance of being proline: the interaction of proline-rich motifs in signaling proteins with their cognate domains. FASEB J. 14, 231241 (2000)
  27. Aitio, O. et al. Recognition of tandem PxxP motifs as a unique Src homology 3-binding mode triggers pathogen-driven actin assembly. Proc. Natl Acad. Sci. USA 107, 2174321748 (2010)
  28. Haapasalo, A. et al. Emerging role of Alzheimer’s disease-associated ubiquilin-1 in protein aggregation. Biochem. Soc. Trans. 38, 150155 (2010)
  29. Kim, S. H. et al. Potentiation of amyotrophic lateral sclerosis (ALS)-associated TDP-43 aggregation by the proteasome-targeting factor, ubiquilin 1. J. Biol. Chem. 284, 80838092 (2009)
  30. Aguzzi, A. & O'Connor, T. Protein aggregation diseases: pathogenicity and therapeutic perspectives. Nature Rev. Drug Discov. 9, 237248 (2010)
  31. Brooks, B. R., Miller, R. G., Swash, M. & Munsat, T. L. El Escorial revisited: revised criteria for the diagnosis of amyotrophic lateral sclerosis. Amyotroph. Lateral Scler. Other Motor Neuron Disord. 1, 293299 (2000)
  32. Neary, D. et al. Frontotemporal lobar degeneration: a consensus on clinical diagnostic criteria. Neurology 51, 15461554 (1998)
  33. Cairns, N. J. et al. Neuropathologic diagnostic and nosologic criteria for frontotemporal lobar degeneration: consensus of the consortium for frontotemporal lobar degeneration. Acta Neuropathol. 114, 522 (2007)
  34. Mackenzie, I. R. et al. Heterogeneity of ubiquitin pathology in frontotemporal lobar degeneration: classification and relation to clinical phenotype. Acta Neuropathol. 112, 539549 (2006)
  35. Deng, H. X. et al. Conversion to the amyotrophic lateral sclerosis phenotype is associated with intermolecular linked insoluble aggregates of SOD1 in mitochondria. Proc. Natl Acad. Sci. USA 103, 71427147 (2006)
  36. Fecto, F. et al. Mutant TRPV4-mediated toxicity is linked to increased constitutive function in axonal neuropathies. J. Biol. Chem. 286, 1728117291 (2011)

Download references

Author information

  1. These authors contributed equally to this work.

    • Han-Xiang Deng &
    • Wenjie Chen


  1. Division of Neuromuscular Medicine, Davee Department of Neurology and Clinical Neurosciences, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA

    • Han-Xiang Deng,
    • Wenjie Chen,
    • Seong-Tshool Hong,
    • George H. Gorrie,
    • Nailah Siddique,
    • Yi Yang,
    • Faisal Fecto,
    • Yong Shi,
    • Hong Zhai,
    • Hujun Jiang,
    • Makito Hirano,
    • Sandra Donkervoort,
    • Kaouther Ajroud,
    • Robert L. Sufit &
    • Teepu Siddique
  2. Department of Pediatrics, University of Ottawa and Children’s Hospital of Eastern Ontario Research Institute, Ottawa, Ontario K1H 8L1, Canada

    • Kym M. Boycott
  3. Interdepartmental Neuroscience Program, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA

    • Faisal Fecto,
    • Enrico Mugnaini &
    • Teepu Siddique
  4. John P. Hussman Institute for Human Genomics, University of Miami, Miller School of Medicine, Miami, Florida 33136, USA

    • Evadnie Rampersaud &
    • Margaret A. Pericak-Vance
  5. Division of Anatomic Pathology, The Ottawa Hospital, Ottawa, Ontario K1Y 4E9, Canada

    • Gerard H. Jansen
  6. Division of Neuropathology, Department of Pathology, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA

    • Eileen H. Bigio
  7. Department of Neurology, Neuroscience and Spine Institute, Carolinas Medical Center, Charlotte, North Carolina 28207, USA

    • Benjamin R. Brooks
  8. Center for Human Genetics Research, Vanderbilt University, Nashville, Tennessee 37232, USA

    • Jonathan L. Haines
  9. Department of Cell and Molecular Biology, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA

    • Enrico Mugnaini &
    • Teepu Siddique
  10. Present addresses: Laboratory of Genetics and Department of Microbiology, Chonbuk National University Medical School, Chonbuk 561-712, South Korea (S.-T.H.); Institute of Neurological Sciences, Southern General Hospital, Glasgow G51 4TF, UK (G.H.G.); Department of Health Sciences, National Natural Science Foundation of China, Beijing 100085, China (H.J.); Department of Neurology, Sakai Hospital Kinki University Faculty of Medicine, Osaka 590-0132, Japan (M.H.).

    • Seong-Tshool Hong,
    • George H. Gorrie,
    • Hujun Jiang &
    • Makito Hirano


T.S. conceived and supervised this project. W.C., S.-T.H., Y.Y., H.J., M.H., H.-X.D. and T.S. did the sequencing analysis. S.T.H., E.R., J.L.H., M.P.-V. and T.S. performed linkage analysis. K.M.B., G.H.G., F.F., G.H.J., H.Z., E.H.B., K.A., E.M., H.-X.D. and T.S. performed immunohistochemical, confocal and pathological analysis. F.F., Y.S. and H.-X.D. performed functional analysis. N.S., S.D. and T.S. collected family information and coordinated this study. K.M.B., G.H.J., B.R.B., R.L.S. and T.S. did clinical studies. H.-X.D. and T.S. analysed the data and wrote the paper.

Competing financial interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to:

Author details

Supplementary information

PDF files

  1. Supplementary Information (2.2M)

    The file contains Supplementary Figures 1-18 with legends, Supplementary Tables 1-2 and Supplementary Case Notes.


  1. Report this comment #25976

    Jahahreeh Finley said:

    Very interesting study on ubiquilin2 and the role the ubiquitin proteasome system plays in the pathogenesis of ALS. However, after reading the headlines referencing this study on various online news outlets (e.g. on the website Science Daily, the title of the article reads "Common Cause of All Forms of Amyotrophic Lateral Sclerosis (ALS) Discovered"), a few questions sprung to mind. Are the authors of this study (Deng et al.) implying, as the Science Daily article states, that a dysfunctional ubiquilin2 protein is a common cause of all forms of ALS (both hereditary and sporadic)? If so, for sporadic ALS cases, what causes the ubiquilin2 protein to become dysfunctional in the first place if a genetic mutation is not present?

    I would think that whatever factor/process that leads to the dysfunctionality of ubiquilin2 would be more readily deserving of the word "cause" with respect to the etiology of sporadic ALS as opposed to the presence and detection of a dysfunctional ubiquilin2 protein itself.

  2. Report this comment #26268

    Glen Kisby said:

    Deng and colleagues offer new insight into the role of the ubiquitin-proteasomal pathway (UPP) in the pathogenesis of an inherited form of ALS. Are these observations relevant to the pathogenesis of sporadic forms of ALS? Guam ALS is also characterized by the presence of ubiquinated skein-like inclusions (Oyanagi et al., Acta Neuropathol 88:405-12, 1994) and a strong environmental etiology (Galasko et al., Neurology 58:90-7, 2002). Epidemiological studies by Zhang and colleagues (Acta Neurol Scand 94:51-9, 1996) showed a strong correlation (p<0.00002) between Guam ALS and human exposure to a cycad plant genotoxin (cycasin), the glucoside of methylazoxymethanol (MAM). Rats fed cycad flour develop neuronal ubiquitin aggregates (Shen et al., Ann Neurol 68:70-80, 2010). We showed that greater than 60% of the genes within the UPP are altered by MAM in the brain of young mice (Kisby et al., Neurobiol Dis 19:108-18, 2005), and this genotoxin appears to alter gene expression through a DNA damage-mediated mechanism (Kisby et al., PloS One 6:e20911, 2011). These provocative findings suggest that environmental factors that persistently perturb gene function might be responsible for triggering the UPP dysfunction in one form of sporadic ALS. Future studies are warranted to either validate or refute this hypothesis.

    Glen Kisby, PhD, Western University of Health Sciences, Lebanon, OR.
    Peter Spencer, PhD, FRCPath, Oregon Health & Science University, Portland, OR.

  3. Report this comment #26741

    Jonathan Miller said:

    Has anyone considered the possiblity of misfolded ubiquilin proteins arising from a prion-like process in the absence of genetic mutation?

Subscribe to comments

Additional data