Identification and functional characterization of novel mutations including frameshift mutation in exon 4 of CSF1R in patients with adult-onset leukoencephalopathy with axonal spheroids and pigmented glia

Abstract

ObjectiveAdult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP) is caused by mutations in CSF1R. Pathogenic mutations in exons 12–22 including coding sequence of the tyrosine kinase domain (TKD) of CSF1R were previously identified. We aimed to identify CSF1R mutations in patients who were clinically suspected of having ALSP and to determine the pathogenicity of novel CSF1R variants.MethodsSixty-one patients who fulfilled the diagnostic criteria of ALSP were included in this study. Genetic analysis of CSF1R was performed for all the coding exons. The haploinsufficiency of CSF1R was examined for frameshift mutations by RT-PCR. Ligand-dependent autophosphorylation of CSF1R was examined in cells expressing CSF1R mutants.ResultsWe identified ten variants in CSF1R including two novel frameshift, five novel missense, and two known missense mutations as well as one known missense variant. Eight mutations were located in TKD. One frameshift mutation (p.Pro104LeufsTer8) and one missense variant (p.His362Arg) were located in the extracellular domain. RT-PCR analysis revealed that the frameshift mutation of p.Pro104LeufsTer8 caused nonsense-mediated mRNA decay. Functional assay revealed that none of the mutations within TKD showed autophosphorylation of CSF1R. The p.His362Arg variant located in the extracellular domain showed comparable autophosphorylation of CSF1R to the wild type, suggesting that this variant is not likely pathogenic.ConclusionsThe detection of the CSF1R mutation outside of the region-encoding TKD may extend the genetic spectrum of ALSP with CSF1R mutations. Mutational analysis of all the coding exons of CSF1R should be considered for patients clinically suspected of having ALSP.Electronic supplementary materialThe online version of this article (10.1007/s00415-018-9017-2) contains supplementary material, which is available to authorized users.

A Novel Splicing Mutation in the CSF1R Gene in a Family With Hereditary Diffuse Leukoencephalopathy With Axonal Spheroids

“…The normal function of CSF1R is essential for proliferation, survival, proliferation, and differentiation of mononuclear phagocytic cells, including microglia in the central nervous system (Stabile et al., 2016). To date, more than 60 genetic loci mutations have been linked to HDLS (Figure 4D) (Guerreiro et al., 2013; Konno et al., 2017; Lynch et al., 2017; Adams et al., 2018; Miura et al., 2018). Almost all of the mutations in CSF1R associated with HDLS are located in the tyrosine kinase domain of the CSF1R protein, encoded by exons 12–21 of the genes (Lynch et al., 2017).…”

A Novel Hypomorphic CSF1R Gene Mutation in the Biallelic State Leading to Fatal Childhood Neurodegeneration

“…9 The p. (Thr833Met) observed in our family was further evaluated by transfecting HEK293T cell lines followed by immunoblot analysis and functional test of CSF1R ligand dependent autophosphorylation (LDA) at important tyrosine residues. 10 This analysis demonstrated that the total CSF1R protein expression levels were normal. But the autophosphorylation was completely lost at the p.Ty546, and p.Tyr708 positions, while the percentage of autophosphorylation (LDA) at p.Tyr723 was 22% of wild-type levels (►Figs.…”

“…Heterozygosity for ALSP causative mutation p.P104Lfs à 8 (exon 4, much before the TKD) causes haploinsufficiency due to nonsense mediated decay of the mRNA from mutant allele. 10 In contrast, the mutation p.Y886Qfs à 55 (exon 21, TKD) causes truncated protein with absent autophosphorylation and causes dominant negative mechanism by forming heterodimers with wild-type chain. 10 Gain-of-function mutations such as K584E, R802V, T633 (gatekeeper) residue mutations led to constitutive activation (ligand independent autophosphor-ylation) and subsequent rapid degradation ultimately leading to decreased protein levels (haploinsufficiency).…”

“…10 In contrast, the mutation p.Y886Qfs à 55 (exon 21, TKD) causes truncated protein with absent autophosphorylation and causes dominant negative mechanism by forming heterodimers with wild-type chain. 10 Gain-of-function mutations such as K584E, R802V, T633 (gatekeeper) residue mutations led to constitutive activation (ligand independent autophosphor-ylation) and subsequent rapid degradation ultimately leading to decreased protein levels (haploinsufficiency). 11 Homozygosity for amorphic alleles (e.g., c.1754-1G > C, p. Y540 à ) have been reported to lead to perinatal death but in these families, the carriers (parents and/or grandparents) were not old enough to have excluded presentation as ALSP later in life.…”

Leucodistrofias e leucoencefalopatias genéticas em adultos: caracterização clínica, molecular e de neuroimagem

“…As alterações da RM de crânio podem anteceder os primeiros sinais clínicos em pelo menos 5 anos (Konno et al, 2014). Outro achado significativo é a presença de pequenas calcificações, predominantemente em lobos frontais, próximas aos cornos anteriores dos ventrículos laterais (Konno et al, 2014 (Miura et al, 2018). Todas as variantes patogênicas levam a formação de proteínas truncadas e à haploinsuficiência de CSF1R, ocasionando a doença (Stabile et al, 2016).…”