Loss-of-function mutations in ATP6V0A2 impair vesicular trafficking, tropoelastin secretion and cell survival

Abstract

Autosomal recessive cutis laxa type 2 (ARCL2), a syndrome of growth and developmental delay and redundant, inelastic skin, is caused by mutations in the a2 subunit of the vesicular ATPase H+-pump (ATP6V0A2). The goal of this study was to define the disease mechanisms that lead to connective tissue lesions in ARCL2. In a new cohort of 17 patients, DNA sequencing of ATP6V0A2 detected either homozygous or compound heterozygous mutations. Considerable allelic and phenotypic heterogeneity was observed, with a missense mutation of a moderately conserved residue p.P87L leading to unusually mild disease. Abnormal N- and/or mucin type O-glycosylation was observed in all patients tested. Premature stop codon mutations led to decreased ATP6V0A2 mRNA levels by destabilizing the mutant mRNA via the nonsense-mediated decay pathway. Loss of ATP6V0A2 either by siRNA knockdown or in ARCL2 cells resulted in distended Golgi cisternae, accumulation of abnormal lysosomes and multivesicular bodies. Immunostaining of ARCL2 cells showed the accumulation of tropoelastin (TE) in the Golgi and in large, abnormal intracellular and extracellular aggregates. Pulse-chase studies confirmed impaired secretion and increased intracellular retention of TE, and insoluble elastin assays showed significantly reduced extracellular deposition of mature elastin. Fibrillin-1 microfibril assembly and secreted lysyl oxidase activity were normal in ARCL2 cells. TUNEL staining demonstrated increased rates of apoptosis in ARCL2 cell cultures. We conclude that loss-of-function mutations in ATP6V0A2 lead to TE aggregation in the Golgi, impaired clearance of TE aggregates and increased apoptosis of elastogenic cells.

Clinical and biochemical features guiding the diagnostics in neurometabolic cutis laxa

“…4,12,13,17,18,24,25,33 PCR products were Figure 2 Diagnostic flowchart for evaluation of a new CL patient with suspected autosomal recessive inheritance (non-type 1). The facial features of PYCR1, ALDH18A1 and GORAB-related syndromes are similar to each other and different from the ATP6V0A2-related ARCL2A.…”

“…As our first description of a novel CL syndrome with variable cortical anomalies and a genetic defect in ATP6V0A2, several new patients have been reported. 12,16 In children diagnosed with ATP6V0A2 mutations, the presence of cobblestone-like brain dysgenesis in combination with glycosylation abnormalities is pathognomonic; the cerebral abnormalities are dominated by bilateral cortical malformation of the posterior parts of the frontal lobes or the temporal lobes and enlarged perivascular Clinical and biochemical features in CL T Gardeitchik et al spaces. Surprisingly, one of the patients (patient 6) with a confirmed PYCR1 mutation also had gyration anomalies: polymicrogyria, a recognizably different type than the cobblestone brain-like abnormalities and obviously, without glycosylation abnormalities.…”

“…Although the presentation of CCA is nonspecific, the combination of a CL syndrome with progeroid features and corpus callosum dysgenesis is highly suggestive. 12,17,20 So far, no brain anomalies have been reported in the patients diagnosed with MACS syndrome. The finding of periventricular cystic lesions in our patient (patient 1) diagnosed with a RIN2 mutation could be a coincidence, however, no suggestive perinatal history or vascular aberrations could be confirmed as an underlying basis of this feature.…”

“…ARCL2A (including ATP6V0A2-CDG, Debré-type CL and wrinkly skin syndrome) is caused by mutations in the ATP6V0A2 gene. 2,12,13 Generalized CL is already present at birth and intriguingly improves over time. Patients have suggestive facial features with hanging eyelids, down-slanting palpebral fissures, muscle hypotonia, epilepsy and mild-to-moderate developmental delay.…”

“…Patients have suggestive facial features with hanging eyelids, down-slanting palpebral fissures, muscle hypotonia, epilepsy and mild-to-moderate developmental delay. 2,12,14 The combination of the biochemical marker of abnormal protein glycosylation (congenital disorder of glycosylation (CDGII)) and cobblestone-like brain dysgenesis in ARCL2 is pathognomonic for ARCL2A. 2,15,16 The recently defined ARCL2B is caused by mutations in the PYCR1 gene, involved in de novo mitochondrial proline synthesis, leading to a progeroid appearance, parchment-like skin, triangular face, joint luxations, muscle hypotonia, dystonic posturing and developmental delay.…”