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A SEL1L mutation links a canine progressive early-onset cerebellar ataxia to the endoplasmic reticulum-associated protein degradation (ERAD) machinery.

Kyöstilä K, Cizinauskas S, Seppälä EH, Suhonen E, Jeserevics J, Sukura A, Syrjä P, Lohi H - PLoS Genet. (2012)

Bottom Line: Pathological and histological examinations indicated cerebellum-restricted neurodegeneration.The mutation segregated fully in the recessive pedigree, and a 10% carrier frequency was indicated in a population cohort.Furthermore, our results have enabled the development of a genetic test for breeders.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, University of Helsinki, Helsinki, Finland.

ABSTRACT
Inherited ataxias are characterized by degeneration of the cerebellar structures, which results in progressive motor incoordination. Hereditary ataxias occur in many species, including humans and dogs. Several mutations have been found in humans, but the genetic background has remained elusive in dogs. The Finnish Hound suffers from an early-onset progressive cerebellar ataxia. We have performed clinical, pathological, and genetic studies to describe the disease phenotype and to identify its genetic cause. Neurological examinations on ten affected dogs revealed rapidly progressing generalized cerebellar ataxia, tremors, and failure to thrive. Clinical signs were present by the age of 3 months, and cerebellar shrinkage was detectable through MRI. Pathological and histological examinations indicated cerebellum-restricted neurodegeneration. Marked loss of Purkinje cells was detected in the cerebellar cortex with secondary changes in other cortical layers. A genome-wide association study in a cohort of 31 dogs mapped the ataxia gene to a 1.5 Mb locus on canine chromosome 8 (p(raw) = 1.1x10(-7), p(genome) = 7.5x10(-4)). Sequencing of a functional candidate gene, sel-1 suppressor of lin-12-like (SEL1L), revealed a homozygous missense mutation, c.1972T>C; p.Ser658Pro, in a highly conserved protein domain. The mutation segregated fully in the recessive pedigree, and a 10% carrier frequency was indicated in a population cohort. SEL1L is a component of the endoplasmic reticulum (ER)-associated protein degradation (ERAD) machinery and has not been previously associated to inherited ataxias. Dysfunctional protein degradation is known to cause ER stress, and we found a significant increase in expression of nine ER stress responsive genes in the cerebellar cortex of affected dogs, supporting the pathogenicity of the mutation. Our study describes the first early-onset neurodegenerative ataxia mutation in dogs, establishes an ERAD-mediated neurodegenerative disease model, and proposes SEL1L as a new candidate gene in progressive childhood ataxias. Furthermore, our results have enabled the development of a genetic test for breeders.

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Histological findings within the cerebellar cortex of affected dogs.(A) Normal cerebellar cortex in the ventrolateral parts of the cerebellum, with densely cellular granular cell layer (HE 100×). (B) Affected cerebellar cortex in the vermal region with marked loss of granular cells (HE 100×). (C) A higher magnification of unaffected cortex with viable Purkinje cells (PCs) and a normal granular cell density (HE 400×). (D) A higher magnification of affected cortex, with severe loss of PCs and linear reactive gliosis between the molecular and the granular layers (arrow) (HE 400×). (E) Degenerating, shrunken, eosinophilic PCs with margination of the nuclear chromatin (black arrow) or central chromatolysis of the cytoplasma (red arrow) (HE 400×). (F) PCs that show total loss of cytoplasmic basophilia (Nissl substance) and pyknotic or karyorhektic nuclei (arrows). Inset: viable PC with intact Nissl substance seen as basophilic cytoplasmic granulation (LFB-CEV, 400×). (G) Left: mild astrogliosis (arrows) within the granular cell layer and the white matter of the cerebellar folia. Right: unaffected part of same dog (IHC GFAP, 400×). (H) Left: secondary degeneration and myelinophagia in the affected cerebellar white matter. Right: unaffected parts of same dog (LFB-CEV, 400×). ML = molecular layer, PL = Purkinje cell layer, GL = granular cell layer, WM = white matter.
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pgen-1002759-g002: Histological findings within the cerebellar cortex of affected dogs.(A) Normal cerebellar cortex in the ventrolateral parts of the cerebellum, with densely cellular granular cell layer (HE 100×). (B) Affected cerebellar cortex in the vermal region with marked loss of granular cells (HE 100×). (C) A higher magnification of unaffected cortex with viable Purkinje cells (PCs) and a normal granular cell density (HE 400×). (D) A higher magnification of affected cortex, with severe loss of PCs and linear reactive gliosis between the molecular and the granular layers (arrow) (HE 400×). (E) Degenerating, shrunken, eosinophilic PCs with margination of the nuclear chromatin (black arrow) or central chromatolysis of the cytoplasma (red arrow) (HE 400×). (F) PCs that show total loss of cytoplasmic basophilia (Nissl substance) and pyknotic or karyorhektic nuclei (arrows). Inset: viable PC with intact Nissl substance seen as basophilic cytoplasmic granulation (LFB-CEV, 400×). (G) Left: mild astrogliosis (arrows) within the granular cell layer and the white matter of the cerebellar folia. Right: unaffected part of same dog (IHC GFAP, 400×). (H) Left: secondary degeneration and myelinophagia in the affected cerebellar white matter. Right: unaffected parts of same dog (LFB-CEV, 400×). ML = molecular layer, PL = Purkinje cell layer, GL = granular cell layer, WM = white matter.

Mentions: Histological changes of the nervous system were restricted to the cerebellum in all examined puppies. The cerebellar cortex showed marked premature degeneration and loss of PCs with consequent neuronal depletion in the granular cell layer (Figure 2B and 2D). The cerebellar vermis and the paramedian lobule were consistently the most severely affected areas. The cranial regions of the cerebellar cortex were more affected than the caudal regions. The ventrolateral parts, including paraflocculi and flocculus, were spared and partially normal (Figure 2A and 2C). In the cortical areas, where severe PC loss was present, glial cells (Bergmans glia) were proliferating between the molecular and the granular cell layers (Figure 2D). The remaining PCs were shrunken and eosinophilic with marginated nuclear chromatin or showed total loss of cytoplasmic basophilic Nissl substance (chromatolysis) (Figure 2E and 2F). The granular cell layer was markedly depleted of neurons and showed mild astrocytosis in areas of profound PCs loss (Figure 2G). Occasional degenerated and vacuolated axons were detected in the granular layer. Mild to moderate ongoing degeneration and myelinophagia was seen within the cerebellar white matter of the severely affected areas (Figure 2H). Neither transsynaptic degeneration in the cerebellar nuclei nor retrograde degeneration of the olivary nucleus was found. Immunohistochemical (IHC) staining for canine distemper virus and parvovirus showed no positivity. The overall severity of the histopathological findings, including active PC degeneration, total granule cell and PC loss, consecutive white matter lesions and the extent of the lesions, are summarized in Table 1.


A SEL1L mutation links a canine progressive early-onset cerebellar ataxia to the endoplasmic reticulum-associated protein degradation (ERAD) machinery.

Kyöstilä K, Cizinauskas S, Seppälä EH, Suhonen E, Jeserevics J, Sukura A, Syrjä P, Lohi H - PLoS Genet. (2012)

Histological findings within the cerebellar cortex of affected dogs.(A) Normal cerebellar cortex in the ventrolateral parts of the cerebellum, with densely cellular granular cell layer (HE 100×). (B) Affected cerebellar cortex in the vermal region with marked loss of granular cells (HE 100×). (C) A higher magnification of unaffected cortex with viable Purkinje cells (PCs) and a normal granular cell density (HE 400×). (D) A higher magnification of affected cortex, with severe loss of PCs and linear reactive gliosis between the molecular and the granular layers (arrow) (HE 400×). (E) Degenerating, shrunken, eosinophilic PCs with margination of the nuclear chromatin (black arrow) or central chromatolysis of the cytoplasma (red arrow) (HE 400×). (F) PCs that show total loss of cytoplasmic basophilia (Nissl substance) and pyknotic or karyorhektic nuclei (arrows). Inset: viable PC with intact Nissl substance seen as basophilic cytoplasmic granulation (LFB-CEV, 400×). (G) Left: mild astrogliosis (arrows) within the granular cell layer and the white matter of the cerebellar folia. Right: unaffected part of same dog (IHC GFAP, 400×). (H) Left: secondary degeneration and myelinophagia in the affected cerebellar white matter. Right: unaffected parts of same dog (LFB-CEV, 400×). ML = molecular layer, PL = Purkinje cell layer, GL = granular cell layer, WM = white matter.
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pgen-1002759-g002: Histological findings within the cerebellar cortex of affected dogs.(A) Normal cerebellar cortex in the ventrolateral parts of the cerebellum, with densely cellular granular cell layer (HE 100×). (B) Affected cerebellar cortex in the vermal region with marked loss of granular cells (HE 100×). (C) A higher magnification of unaffected cortex with viable Purkinje cells (PCs) and a normal granular cell density (HE 400×). (D) A higher magnification of affected cortex, with severe loss of PCs and linear reactive gliosis between the molecular and the granular layers (arrow) (HE 400×). (E) Degenerating, shrunken, eosinophilic PCs with margination of the nuclear chromatin (black arrow) or central chromatolysis of the cytoplasma (red arrow) (HE 400×). (F) PCs that show total loss of cytoplasmic basophilia (Nissl substance) and pyknotic or karyorhektic nuclei (arrows). Inset: viable PC with intact Nissl substance seen as basophilic cytoplasmic granulation (LFB-CEV, 400×). (G) Left: mild astrogliosis (arrows) within the granular cell layer and the white matter of the cerebellar folia. Right: unaffected part of same dog (IHC GFAP, 400×). (H) Left: secondary degeneration and myelinophagia in the affected cerebellar white matter. Right: unaffected parts of same dog (LFB-CEV, 400×). ML = molecular layer, PL = Purkinje cell layer, GL = granular cell layer, WM = white matter.
Mentions: Histological changes of the nervous system were restricted to the cerebellum in all examined puppies. The cerebellar cortex showed marked premature degeneration and loss of PCs with consequent neuronal depletion in the granular cell layer (Figure 2B and 2D). The cerebellar vermis and the paramedian lobule were consistently the most severely affected areas. The cranial regions of the cerebellar cortex were more affected than the caudal regions. The ventrolateral parts, including paraflocculi and flocculus, were spared and partially normal (Figure 2A and 2C). In the cortical areas, where severe PC loss was present, glial cells (Bergmans glia) were proliferating between the molecular and the granular cell layers (Figure 2D). The remaining PCs were shrunken and eosinophilic with marginated nuclear chromatin or showed total loss of cytoplasmic basophilic Nissl substance (chromatolysis) (Figure 2E and 2F). The granular cell layer was markedly depleted of neurons and showed mild astrocytosis in areas of profound PCs loss (Figure 2G). Occasional degenerated and vacuolated axons were detected in the granular layer. Mild to moderate ongoing degeneration and myelinophagia was seen within the cerebellar white matter of the severely affected areas (Figure 2H). Neither transsynaptic degeneration in the cerebellar nuclei nor retrograde degeneration of the olivary nucleus was found. Immunohistochemical (IHC) staining for canine distemper virus and parvovirus showed no positivity. The overall severity of the histopathological findings, including active PC degeneration, total granule cell and PC loss, consecutive white matter lesions and the extent of the lesions, are summarized in Table 1.

Bottom Line: Pathological and histological examinations indicated cerebellum-restricted neurodegeneration.The mutation segregated fully in the recessive pedigree, and a 10% carrier frequency was indicated in a population cohort.Furthermore, our results have enabled the development of a genetic test for breeders.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, University of Helsinki, Helsinki, Finland.

ABSTRACT
Inherited ataxias are characterized by degeneration of the cerebellar structures, which results in progressive motor incoordination. Hereditary ataxias occur in many species, including humans and dogs. Several mutations have been found in humans, but the genetic background has remained elusive in dogs. The Finnish Hound suffers from an early-onset progressive cerebellar ataxia. We have performed clinical, pathological, and genetic studies to describe the disease phenotype and to identify its genetic cause. Neurological examinations on ten affected dogs revealed rapidly progressing generalized cerebellar ataxia, tremors, and failure to thrive. Clinical signs were present by the age of 3 months, and cerebellar shrinkage was detectable through MRI. Pathological and histological examinations indicated cerebellum-restricted neurodegeneration. Marked loss of Purkinje cells was detected in the cerebellar cortex with secondary changes in other cortical layers. A genome-wide association study in a cohort of 31 dogs mapped the ataxia gene to a 1.5 Mb locus on canine chromosome 8 (p(raw) = 1.1x10(-7), p(genome) = 7.5x10(-4)). Sequencing of a functional candidate gene, sel-1 suppressor of lin-12-like (SEL1L), revealed a homozygous missense mutation, c.1972T>C; p.Ser658Pro, in a highly conserved protein domain. The mutation segregated fully in the recessive pedigree, and a 10% carrier frequency was indicated in a population cohort. SEL1L is a component of the endoplasmic reticulum (ER)-associated protein degradation (ERAD) machinery and has not been previously associated to inherited ataxias. Dysfunctional protein degradation is known to cause ER stress, and we found a significant increase in expression of nine ER stress responsive genes in the cerebellar cortex of affected dogs, supporting the pathogenicity of the mutation. Our study describes the first early-onset neurodegenerative ataxia mutation in dogs, establishes an ERAD-mediated neurodegenerative disease model, and proposes SEL1L as a new candidate gene in progressive childhood ataxias. Furthermore, our results have enabled the development of a genetic test for breeders.

Show MeSH
Related in: MedlinePlus