Congenital disorder of glycosylation caused by starting site-specific variant in syntaxin-5

Peter T A Linders,Kai Muru,Fokje Zijlstra,Sander Pajusalu,Richard Arts,Geert Van Den Bogaart,Karin Huijben,Olga Fjodorova,Mari-Anne Vals,Katrin Õunap,Martin Ter Beest,Kimiyo Raymond,Melissa Baerenfaenger,Dirk Lefeber,Eveline C F Gerretsen,Natalia H Revelo,Angel Ashikov,Rinse De Boer

doi:10.1038/s41467-021-26534-y

Abstract

The SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein syntaxin-5 (Stx5) is essential for Golgi transport. In humans, the STX5 mRNA encodes two protein isoforms, Stx5 Long (Stx5L) from the first starting methionine and Stx5 Short (Stx5S) from an alternative starting methionine at position 55. In this study, we identify a human disorder caused by a single missense substitution in the second starting methionine (p.M55V), resulting in complete loss of the short isoform. Patients suffer from an early fatal multisystem disease, including severe liver disease, skeletal abnormalities and abnormal glycosylation. Primary human dermal fibroblasts isolated from these patients show defective glycosylation, altered Golgi morphology as measured by electron microscopy, mislocalization of glycosyltransferases, and compromised ER-Golgi trafficking. Measurements of cognate binding SNAREs, based on biotin-synchronizable forms of Stx5 (the RUSH system) and Förster resonance energy transfer (FRET), revealed that the short isoform of Stx5 is essential for intra-Golgi transport. Alternative starting codons of Stx5 are thus linked to human disease, demonstrating that the site of translation initiation is an important new layer of regulating protein trafficking.

Highlights

IntroductionThe SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein syntaxin-5 (Stx5) is essential for Golgi transport
The SNARE protein syntaxin-5 (Stx5) is essential for Golgi transport
We demonstrate that Stx5 Long (Stx5L) can largely compensate for the lack of Stx5 Short (Stx5S), the loss of Stx5S leads to defects in intra-Golgi trafficking with altered morphology of the endoplasmic reticulum (ER) and Golgi apparatus and mislocalization of glycosyltransferases, which results in pronounced defects in glycosylation

Summary

Introduction

The SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein syntaxin-5 (Stx5) is essential for Golgi transport. We identified a genetic variant in the second translation codon methionine-55, fully abrogating the production of Stx5S and providing an opportunity to study the physiological relevance of the existence of two isoforms in humans Patients homozygous for this mutation have a severe clinical phenotype associated with infantile mortality and defective protein glycosylation. A mutation in an alternate starting site of ribosomal translation of Stx[5] is linked to human disease This finding reveals that protein function can be regulated at the level of translation initiation and has profound effects on intracellular membrane trafficking and Golgi function

Methods

Results

Conclusion