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Refining genotype-phenotype correlation in Alstrom syndrome through study of primary human fibroblasts.

ABSTRACT: Alström syndrome (AS), featuring retinal dystrophy, neuronal deafness, cardiomyopathy, metabolic syndrome, and diffuse fibrosis, is caused by biallelic mutations in the centrosomal protein ALMS1. Genotype-phenotype correlation has been suggested without assessment of ALMS1 expression.ALMS1 expression (real-time PCR and immunocytochemistry) and cilia formation (immunocytochemistry) were assessed in fibroblasts from deeply phenotyped volunteers diagnosed with AS recruited from a dedicated AS Service. Exome sequencing was used in two participants without convincing biallelic ALMS1 mutations, and BBS2 (Bardet-Biedl syndrome 2) protein expression was assessed in one patient with biallelic BBS2 mutations. Hedgehog-induced GLI1 expression and PDGFA signaling was assessed using quantitative real-time PCR, immunoblotting, or immunostaining of fixed cells after stimulation.In 16 of the patient cell lines examined, ALMS1 protein was undetectable (14 with biallelic loss-of-function (LoF) mutations), and in two, ALMS1 staining was equivocal (one with biallelic LoF mutations). In five lines, ALMS1 expression was normal using at least one fixation method (one with biallelic LoF mutations). These differences were not accounted for by major differences in ALMS1 mRNA expression. Exome sequencing of two participants with normal ALMS1 expression identified biallelic LoF BBS2 mutations in one. No second, known ciliopathy mutation was found in the other patient, who had one LoF ALMS1 mutation. Phenotypes were milder or atypical in participants with preserved ALMS1 immunostaining, even when two with likely alternative genetic diagnoses were excluded. All cells studied developed normal cilia, ALMS1 and BBS2 mutant cells showed normal Hedgehog-induced upregulation of GLI1 expression, and PDGFA signaling was normal in ALMS1-deficient cells.Milder or atypical presentations of AS should prompt genetic evaluation for alternative, clinically overlapping ciliopathies. A subgroup of patients with bona fide ALMS1 defects have milder phenotypes due to residual ALMS1 expression, which may be more important than mutation site.

SUBMITTER: Chen JH

PROVIDER: S-EPMC5511801 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Refining genotype-phenotype correlation in Alström syndrome through study of primary human fibroblasts.

Chen Jian-Hua JH Geberhiwot Tarekegn T Barrett Timothy G TG Paisey Richard R Semple Robert K RK

Molecular genetics & genomic medicine 20170515 4

<h4>Background</h4>Alström syndrome (AS), featuring retinal dystrophy, neuronal deafness, cardiomyopathy, metabolic syndrome, and diffuse fibrosis, is caused by biallelic mutations in the centrosomal protein ALMS1. Genotype-phenotype correlation has been suggested without assessment of ALMS1 expression.<h4>Methods</h4>ALMS1 expression (real-time PCR and immunocytochemistry) and cilia formation (immunocytochemistry) were assessed in fibroblasts from deeply phenotyped volunteers diagnosed with AS ...[more]

PMID: 28717663

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Project description:Betaine-homocysteine methyltransferase (BHMT) catalyzes the remethylation of homocysteine. BHMT is highly expressed in the human liver. In the liver, BHMT catalyzes up to 50% of homocysteine metabolism. Understanding the relationship between BHMT genetic polymorphisms and function might increase our understanding of the role of this reaction in homocysteine remethylation and in S-adenosylmethionine-dependent methylation. To help achieve those goals, we measured levels of BHMT enzyme activity and immunoreactive protein in 268 human hepatic surgical biopsy samples from adult subjects as well as 73 fetal hepatic tissue samples obtained at different gestational ages. BHMT protein levels were correlated significantly (p<0.001) with levels of enzyme activity in both fetal and adult tissues, but both were decreased in fetal tissue when compared with levels in the adult hepatic biopsies. To determine possible genotype-phenotype correlations, 12 tag SNPs for BHMT and the closely related BHMT2 gene were selected from SNPs observed during our own gene resequencing studies as well as from HapMap. These SNPs data were used to genotype DNA from the adult hepatic surgical biopsy samples, and genotype-phenotype association analysis was performed. Three SNPs (rs41272270, rs16876512, and rs6875201), located 28kb upstream, in the 5'-UTR and in intron 1 of BHMT, respectively, were significantly correlated with both BHMT activity (p=3.41E-8, 2.55E-9 and 2.46E-10, respectively) and protein levels (p=5.78E-5, 1.08E-5 and 6.92E-6, respectively). We also imputed 230 additional SNPs across the BHMT and BHMT2 genes, identifying an additional imputed SNP, rs7700790, that was also highly associated with hepatic BHMT enzyme activity and protein. However, none of the 3 genotyped or one imputed SNPs displayed a "shift" during electrophoretic mobility shift assays. These observations may help us to understand individual variation in the regulation of BHMT in the human liver and its possible relationship to variation in methylation.

Dataset Information

Refining genotype-phenotype correlation in Alstrom syndrome through study of primary human fibroblasts.

Publications

Refining genotype-phenotype correlation in Alström syndrome through study of primary human fibroblasts.

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