Translation initiator EIF4G1 mutations in familial Parkinson disease

Abstract

Genome-wide analysis of a multi-incident family with autosomal-dominant parkinsonism has implicated a locus on chromosomal region 3q26-q28. Linkage and disease segregation is explained by a missense mutation c.3614G>A (p.Arg1205His) in eukaryotic translation initiation factor 4-gamma (EIF4G1). Subsequent sequence and genotype analysis identified EIF4G1 c.1505C>T (p.Ala502Val), c.2056G>T (p.Gly686Cys), c.3490A>C (p.Ser1164Arg), c.3589C>T (p.Arg1197Trp) and c.3614G>A (p.Arg1205His) substitutions in affected subjects with familial parkinsonism and idiopathic Lewy body disease but not in control subjects. Despite different countries of origin, persons with EIF4G1 c.1505C>T (p.Ala502Val) or c.3614G>A (p.Arg1205His) mutations appear to share haplotypes consistent with ancestral founders. eIF4G1 p.Ala502Val and p.Arg1205His disrupt eIF4E or eIF3e binding, although the wild-type protein does not, and render mutant cells more vulnerable to reactive oxidative species. EIF4G1 mutations implicate mRNA translation initiation in familial parkinsonism and highlight a convergent pathway for monogenic, toxin and perhaps virally-induced Parkinson disease.

Figure 1

Pedigrees with EIF4G1 Mutations and Parkinsonism Individual pedigrees are numbered and their country of origin is indicated. Only pedigrees with an (A) EIF4G1 c.3614G>A (p.Arg1205His), (B) c.1505C>T (p.Ala502Val), (C) c.2056G>T (p.Gly686Cys), (D) c.2056G>T (p.Gly686Cys) and c.3589C>T (p.Arg1197Trp) and (E) c.3490A>C (p.Ser1164Arg) mutations are illustrated. Filled symbols indicate affected individuals with parkinsonism; the age of symptom onset is shown in parentheses. Quarter-filled symbols indicate individuals with dementia. To protect confidentiality, the pedigrees do not show some individuals and siblings and/or gender is sometimes disguised with a diamond. All samples genotyped are indicted as heterozygous mutations (M) or as wild-type (wt). The asterisk (^∗) indicate that this individual did not fulfill the criteria for PD, but presented resting tremor and akinesia.

Figure 2

Disease-Segregating EIF4G1 c.3614G>A Haplotypes among Families (A) Comparison of haplotype data among all families with EIF4G1 mutations shows alleles between rs4912537 and D3S3578 are shared (Table S2). The maximum shared interval is 99,330 bp (MAX; chr3:185,519,706–185,619,035; gray and blue bars) and contains six gene transcripts, EIF4G1, FAM131A, CLCN2, POLR2H, THPO, and CHRD. The minimal shared interval is 45,676 bp (MIN; chr3:185,521,663-185,567,338; gray) and harbors four genes (EIF4G1, FAM131A, CLCN2, and POLR2H). EIF4G1 c.3614G>A (p.Arg1205His) is the only novel coding variant that segregates with disease. (B) The CEU haplotype map suggests negligible linkage disequilibrium across the rs4912537–D3S3578 region.

Figure 3

Coimmunoprecipitation Studies of eIF4G1 p.Ala502Val and p.Arg1205His with eIF4A and eIF3e Coimmunoprecipitation (CO-IP) of endogenous eIF4A, eIF4E, and eIF3e with transfected full-length eIF4G1-V5 (WT, a dominant negative mutant, or the p.Ala502Val and p.Arg1205His mutants) in HEK293T cells. (A) The p.Ala502Val mutant perturbs the interaction with eIF4E but not eIF4A (IP-V5). The inputs on the upper right panel represent the amounts of transfected eIF4G1-V5 (WT and mutants) and the endogenous proteins (eIF4A1 or eIF4E) introduced in the immunoprecipitation assay. (B) Coimmunoprecipitation of eIF3e with eIF4G1-V5 (IP left; inputs right). In the presence of the p.Arg1205His mutation significantly reduced amounts of eIF3e protein are coprecipitated with eIF4G1-V5. Experiments were performed on three separate occasions as depicted in the graphs. Representative immunoblots are shown.

Figure 4

Fluorescence-Activated Cell-Sorting Analysis of Mutant and Control Cell Lines HEK293T Cells Transiently Overexpressing WT or Mutant EIF4G1 (p.Ala502Val, p.Arg1205His) eIF4G1-GFP proteins were subjected to FACS analysis, with TMRE labeling applied as a marker of mitochondrial polarization. Untreated eIF4G1-GFP-positive cells are comparable in terms of TMRE loading. In contrast, with hydrogen peroxide treatment p.Ala502Val- and p.Arg1205His eIF4G1-GFP-positive cells show a 10% decrease in cell survival compared to WT cells. For each sample, 100,000 events were acquired and subsequently analyzed. After the removal of background counts and the exclusion of untransfected (GFP negative) cells, average TMRE loading values were based on at least 25,000 cells. The number of TMRE-positive cells was calculated as a percentage of the total number of GFP-positive cells in the respective sample. Experiments with two duplicates per condition were performed on three separate occasions.