Molecular analysis of a new type of spinocerebellar degeneration with GFAP mutations

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Glial fibrillary acidic protein (GFAP) is an intermediate filament protein expressed in astrocyte. Recently heterozygous point mutations in the GFAP have been reported in patients with various forms of Alexander disease (ALX). ALX is classified into infantile, juvenile and adult forms by age of disease onset. Infantile form is characterized by progressive psychomotor deterioration, seizure, hydrocephalus, and dies in infancy. Juvenile form is characterized by psychomotor deterioration, bulbar signs, and weakness and spasticity of lower limbs, but mental retardation and progression are less severe than those of the infantile form. Adult form is characterized by ataxia, palatal myoclonus and spastic paraparesis, but the symptoms are extremely variable from severe neurological defects to asymptomatic state.We reported familial cases of palatal myoclonus and spinal cord atrophy with V87G GFAP mutation. We also reported a new case of palatal myoclonus, pyramidal tract signs, cerebellar sign … More s, and marked atrophy of the medulla oblongata and spinal cord, and heterozygous R416W mutation which was also found in both of infantile and juvenile forms. To clarify the correlation of the clinical phenotypes and GFAP mutations, we analyzed nine patients in four Japanese families and reviewed 65 reported patients on the point of GFAP mutations. In addition, we produced the mutant GFAP expression vectors and investigated the expression pattern of the mutant GFAP in the culture cells.In the 74 patients, 42, 13 and 19 patients were infantile, juvenile and adult forms, respectively. In the 19 patients with adult form, two of them had no subjective symptoms and six of seven families had family history of the disease. The juvenile and adult forms coexisted in the same family members. Palatal myoclonus was recognized only four patients with the adult form. Regarding MRI findings, frontal dominant white matter lesion was detected in the infantile form, but spinal cord atrophy was prominent in the adult form. The sites of GFAP mutation mainly localized at exon 1, 4 and 8. R416W mutation was detected all forms of ALX and no clear correlation between the clinical forms and nature of the mutations. Mitochondrial abnormalities, ragged-red fibers, mtDNA A8291G substitution and 9bp deletion, were detected in one patient with the juvenile form. No nestin mutation was detected in the adult form.We produced four different GFAP mutations (T235C, T274G, C276T, C1260T) by site specific mutagenesis. These mutants were transfected into cloned human glioma cells. We investigated the expressions properties of the GFAP protein through immunohistochemical staining using anti-GFAP antibody. Over expression of GFAP was found in the cells with GFAP mutations in comparison with those of wild type GFAP.In conclusion, we confirmed the phenotypic and genetic features of adult form ALX with GFAP mutations. The phenotypic difference between each form of ALX is due not only to the different site and nature of mutations in GFAP, but also to other modifying factor(s) like other intermediate filaments and mitochondrial functions. Comparison of the expression patterns of each mutation in transgenic mice and culture system may clarify the phenotypic and genetic correlation in ALX with GFAP mutations. Less