Laforin, defective in the progressive myoclonus epilepsy of Lafora type, is a dual-specificity phosphatase associated with polyribosomes

The progressive myoclonus epilepsy of Lafora type is an autosomal recessive disorder caused by mutations in the EPM2A gene. EPM2A is predicted to encode a putative tyrosine phosphatase protein, named laforin, whose full sequence has not yet been reported. In order to understand the function of the E...

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Published in:Human molecular genetics 2000-09, Vol.9 (15), p.2251-2261
Main Authors: GANESH, Subramaniam, AGARWALA, Kishan Lal, UEDA, Kazunori, AKAGI, Takumi, SHODA, Keiko, USUI, Takeo, HASHIKAWA, Tsutomu, OSADA, Hiroyuki, DELGADO-ESCUETA, Antonio V, YAMAKAWA, Kazuhiro
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Base Sequence
Biological and medical sciences
Cell Fractionation
Cloning, Molecular
DNA, Complementary - analysis
DNA, Complementary - isolation & purification
EPM2A gene
Fluorescent Antibody Technique
Headache. Facial pains. Syncopes. Epilepsia. Intracranial hypertension. Brain oedema. Cerebral palsy
HeLa Cells
Humans
Lafora Disease - genetics
Lafora Disease - metabolism
Medical sciences
Microscopy, Confocal
Microscopy, Electron
Molecular Sequence Data
Mutation, Missense
myoclonus epilepsy of Lafora type
Nervous system (semeiology, syndromes)
Neurology
Polyribosomes - metabolism
Protein Binding
Protein Conformation
Protein Folding
Protein Tyrosine Phosphatases - genetics
Protein Tyrosine Phosphatases - metabolism
Protein Tyrosine Phosphatases, Non-Receptor
Recombinant Fusion Proteins - genetics
Recombinant Fusion Proteins - metabolism
Transfection
Ubiquitins - metabolism
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Summary:The progressive myoclonus epilepsy of Lafora type is an autosomal recessive disorder caused by mutations in the EPM2A gene. EPM2A is predicted to encode a putative tyrosine phosphatase protein, named laforin, whose full sequence has not yet been reported. In order to understand the function of the EPM2A gene, we isolated a full-length cDNA, raised an antibody and characterized its protein product. The full-length clone predicts a 38 kDa laforin that was very close to the size detected in transfected cells. Recombinant laforin was able to hydrolyze phosphotyrosine as well as phosphoserine/threonine substrates, demonstrating that laforin is an active dual-specificity phosphatase. Biochemical, immunofluorescence and electron microscopic studies on the full-length laforin expressed in HeLa cells revealed that laforin is a cytoplasmic protein associated with polyribosomes, possibly through a conformation-dependent protein-protein interaction. We analyzed the intracellular targeting of two laforin mutants with missense mutations. Expression of both mutants resulted in ubiquitin-positive perinuclear aggregates suggesting that they were misfolded proteins targeted for degradation. Our results suggest that laforin is involved in translational regulation and that protein misfolding may be one of the molecular bases of the Lafora disease phenotype caused by missense mutations in the EPM2A gene.
ISSN:0964-6906
1460-2083
DOI:10.1093/oxfordjournals.hmg.a018916