Abnormal glycosylation in Joubert syndrome type 10

Abstract

BackgroundThe discovery of disease pathogenesis requires systematic agnostic screening of multiple homeostatic processes that may become deregulated. We illustrate this principle in the evaluation and diagnosis of a 5-year-old boy with Joubert syndrome type 10 (JBTS10). He carried the OFD1 mutation p.Gln886Lysfs*2 (NM_003611.2: c.2656del) and manifested features of Joubert syndrome.MethodsWe integrated exome sequencing, MALDI-TOF mass spectrometry analyses of plasma and cultured dermal fibroblasts glycomes, and full clinical evaluation of the proband. Analyses of cilia formation and lectin staining were performed by immunofluorescence. Measurement of cellular nucleotide sugar levels was performed with high-performance anion-exchange chromatography with pulsed amperometric detection. Statistical analyses utilized the Student’s and Fisher’s exact t tests.ResultsGlycome analyses of plasma and cultured dermal fibroblasts identified abnormal N- and O-linked glycosylation profiles. These findings replicated in two unrelated males with OFD1 mutations. Cultured fibroblasts from affected individuals had a defect in ciliogenesis. The proband’s fibroblasts also had an abnormally elevated nuclear sialylation signature and increased total cellular levels of CMP-sialic acid. Ciliogenesis and each glycosylation anomaly were rescued by expression of wild-type OFD1.ConclusionsThe rescue of ciliogenesis and glycosylation upon reintroduction of WT OFD1 suggests that both contribute to the pathogenesis of JBTS10.Electronic supplementary materialThe online version of this article (doi:10.1186/s13630-017-0048-6) contains supplementary material, which is available to authorized users.

Mutations in ARMC9, which Encodes a Basal Body Protein, Cause Joubert Syndrome in Humans and Ciliopathy Phenotypes in Zebrafish

“…We cultured human fibroblasts from healthy controls in DMEM þ 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin under standard conditions. [59][60][61] To induce ciliogenesis, we cultured in DMEM without FBS once the cells reached 70% confluency. After treatment with 0.05% trypsin, cycling cells and serumstarved cells were harvested and RNA extracted using the Aurum Total RNA Mini Kit (Bio-rad).…”

Defining Disease, Diagnosis, and Translational Medicine within a Homeostatic Perturbation Paradigm: The National Institutes of Health Undiagnosed Diseases Program Experience

“…Exemplifying the utility of this agnostic approach, approximately 50% of UDP patients screened for perturbation of protein glycosylation or free glycans in the plasma or urine differed from healthy controls (data not shown). These qualitative and quantitative changes in glycosylation, whether primary or secondary, have diagnostic, mechanistic, or therapeutic value as illustrated by detection of glycosylation abnormalities in DNA repair disorders ( 27 , 28 ), ciliopathies ( 29 , 30 ), mitochondriopathies ( 31 , 32 ), and Golgi disorders ( 33 ). In contrast, detailed metabolomics studies uncovered very few anomalies, suggesting that the current medical testing technology already detects most disorders of metabolism prior to referral to the NIH UDP (data not shown).…”

Mechanisms of Impaired Lung Development and Ciliation in Mannosidase-1-Alpha-2 (Man1a2) Mutants

“…Pcm1 and Dgke were upregulated in NAC-treated live WT and heterozygous pups and downregulated in dead heterozygous and null pups. Defective glycosylation of ciliary proteins like Jbt17, which is performed by enzymes such as Man1a2 can lead to ciliopathies (Kane et al, 2017).…”