RESUMO
Following the mutation screening of genes known to cause amyotrophic lateral sclerosis (ALS) in index cases from 107 familial ALS (FALS) kindred, a point mutation was identified in vesicle-associated membrane protein-associated protein B (VAPB), or VAMP-associated protein B, causing an amino acid change from threonine to isoleucine at codon 46 (T46I) in one FALS case but not in 257 controls. This is an important finding because it is only the second mutation identified in this gene that causes ALS. In order to investigate the pathogenic effects of this mutation, we have used a motor neuron cell line and tissue-specific expression of the mutant protein in Drosophila. We provide substantial evidence for the pathogenic effects of this mutation in abolishing the effect of wild type VAPB in the unfolded protein response, promoting ubiquitin aggregate formation, and activating neuronal cell death. We also report that expression of the mutant protein in the Drosophila motor system induces aggregate deposition, endoplasmic reticulum disorganization, and chaperone up-regulation both in neurons and in muscles. Our integrated analysis of the pathogenic effect of the T46I mutation and the previously identified P56S mutation indicate extensive commonalities in the disease mechanism for these two mutations. In summary, we show that this newly identified mutation in human FALS has a pathogenic effect, supporting and reinforcing the role of VAPB as a causative gene of ALS.
Assuntos
Substituição de Aminoácidos , Esclerose Lateral Amiotrófica , Doenças Genéticas Inatas , Proteínas Interatuantes com Canais de Kv , Mutação de Sentido Incorreto , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/patologia , Animais , Animais Geneticamente Modificados , Células COS , Chlorocebus aethiops , Estudos de Coortes , Drosophila melanogaster , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/patologia , Feminino , Regulação da Expressão Gênica/genética , Doenças Genéticas Inatas/genética , Doenças Genéticas Inatas/metabolismo , Doenças Genéticas Inatas/patologia , Chaperonas de Histonas/biossíntese , Chaperonas de Histonas/genética , Humanos , Proteínas Interatuantes com Canais de Kv/genética , Proteínas Interatuantes com Canais de Kv/metabolismo , Masculino , Especificidade de Órgãos/genéticaRESUMO
The aim of this study was to quantify spinal cord expression of genes known to cause familial amyotrophic lateral sclerosis (FALS) or influence survival in a large cohort of sporadic cases of ALS (SALS), in order to determine their relevance to pathogenic mechanisms occurring in SALS. The expression of superoxide dismutase 1 (SOD1), vesicle associated membrane protein (VAPB), senataxin (SETX), dynactin (DCTN1), vascular endothelial growth factor (VEGF), insulin-like growth factor-1 (IGF1), the small heat shock proteins, HSPB1 and HSPB8, and three genes activated during disease progression, caspases-1 and -3 and glial fibrillary acidic protein (GFAP), were quantified. Robust changes in the expression of four genes were found, VAPB mRNA levels were decreased in the spinal cord of ALS patients compared to controls (p<0.006), whilst HSPB1, HSPB8 and caspase-1 showed significant increases (1.5-2.3-fold). Expression of VAPB mRNA and protein was predominantly localised to large motor neurones further supporting the relevance of this finding to disease progression occurring in SALS.
