Total loss of GM3 synthase activity by a normally processed enzyme in a novel variant and in all ST3GAL5 variants reported to cause a distinct congenital disorder of glycosylation

Abstract

ST3GAL5-CDG is a rare syndrome which is caused by variant GM3 synthases, the enzyme involved in the biosynthesis of a-b-c-series gangliosides. Here we report a novel homozygous ST3GAL5 variant, p.Gly342Ser, in a patient suffering from failure to thrive, severe hearing, visual, motor, and cognitive impairment, and respiratory chain dysfunction. A GM3 synthase assay towards the natural acceptor substrate lactosylceramide was performed upon transfection in HEK-293T cells of expression plasmids carrying wild type and mutated ST3GAL5 cDNAs. The assay revealed a complete loss of enzyme activity. Identical results were obtained with the other four ST3GAL5 variants which have been reported to be pathogenic. HEK-293T clones permanently expressing HaloTag-ST3GAL5 carrying each of the five variants were assessed by quantitative PCR, flow cytometry, western blotting, and confocal microscopy. The results indicated that transcription, translation, stability, and intracellular localization of the tagged protein were identical to those of the wild type construct. Compared with the very mild phenotype of st3gal5 KO mouse models, the results suggest that unknown mechanisms, in addition to the lack of a-b-c-series gangliosides, contribute to the syndrome. Direct enzyme assay upon transfection in model cells appears to be an effective tool for characterizing variants of glycosyltransferases involved in glycosphingolipid biosynthesis.

The Link between Gaucher Disease and Parkinson’s Disease Sheds Light on Old and Novel Disorders of Sphingolipid Metabolism

“…Altogether, this data is in agreement with the current hypothesis that the total absence of gangliosides is responsible for the general lack of neuron homeostasis, leading to inflammation and, consequently, loss of several functions [105]. However, many patients present mildly elevated blood lactate and their cells have an impaired respiratory chain function [25,100]. In this context, the hypothesis that impaired autophagy and, in particular, mitophagy may be involved in the disease pathogenesis should be kept in mind.…”

“…Some of these patients lack clinical seizures or were initially able to crawl, as originally reported only for two siblings carrying the compound C195S/G201A mutations [103]. Accordingly, we have recently shown that all reported mutations abolish any detectable enzyme activity, without differences between the single mutations [100]. Moreover, we also found a normal localization of the variant ST3GAL5s.…”

“…Five different pathogenic mutations have been reported so far, all causing a very severe syndrome characterized by normal pregnancy and delivery but early onset of neurological symptoms including drug-resistant epilepsy, failure to thrive, and impaired hearing and vision. Growth delay and progressive regression occur, giving rise to deafness, blindness, and general motor and cognitive impairment [25,100]. While the original description of a few families suggested a potential different degree of pathogenicity between mutations [101,102,103], a recent survey of several cases affected by the same R288* non-sense mutation indicated the opposite [104].…”

Metabolism of Glycosphingolipids and Their Role in the Pathophysiology of Lysosomal Storage Disorders

“…The ganglioside patterns in mice mimic those in a human newborn with SD and TSD, and mouse models have greatly advanced our knowledge of ganglioside metabolism and therapy of LSDs [14]. However, in certain types of LSDs, the symptoms developing in mice were less severe than those in humans suggesting a greater complexity of the human syndrome [15]. For example, the SD and TSD disease models in mice do not completely recapitulate the human phenotype.…”

Neural-specific alterations in glycosphingolipid biosynthesis and cell signaling associated with two human ganglioside GM3 Synthase Deficiency variants

“…The variant associated with Salt & Pepper Syndrome, ST3GAL5 c.1063G>A (p.Glu355Lys), encodes for a mutant form of the enzyme lacking any enzymatic activity [9]. A more prevalent variant identified in the Amish population, c.862C>T (p.Arg288Ter), encodes for a truncated and inactive enzyme [9,10].…”