Cargando…

N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2

The Sd(a) carbohydrate epitope and its biosynthetic B4GALNT2 enzyme are expressed in the healthy colon and down-regulated to variable extents in colon cancer. The human B4GALNT2 gene drives the expression of a long and a short protein isoform (LF-B4GALNT2 and SF-B4GALNT2) sharing identical transmemb...

Descripción completa

Detalles Bibliográficos
Autores principales: Cogez, Virginie, Vicogne, Dorothée, Schulz, Céline, Portier, Lucie, Venturi, Giulia, de Ruyck, Jérôme, Decloquement, Mathieu, Lensink, Marc F., Brysbaert, Guillaume, Dall’Olio, Fabio, Groux-Degroote, Sophie, Harduin-Lepers, Anne
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9959560/
https://www.ncbi.nlm.nih.gov/pubmed/36835549
http://dx.doi.org/10.3390/ijms24044139
_version_ 1784895307371249664
author Cogez, Virginie
Vicogne, Dorothée
Schulz, Céline
Portier, Lucie
Venturi, Giulia
de Ruyck, Jérôme
Decloquement, Mathieu
Lensink, Marc F.
Brysbaert, Guillaume
Dall’Olio, Fabio
Groux-Degroote, Sophie
Harduin-Lepers, Anne
author_facet Cogez, Virginie
Vicogne, Dorothée
Schulz, Céline
Portier, Lucie
Venturi, Giulia
de Ruyck, Jérôme
Decloquement, Mathieu
Lensink, Marc F.
Brysbaert, Guillaume
Dall’Olio, Fabio
Groux-Degroote, Sophie
Harduin-Lepers, Anne
author_sort Cogez, Virginie
collection PubMed
description The Sd(a) carbohydrate epitope and its biosynthetic B4GALNT2 enzyme are expressed in the healthy colon and down-regulated to variable extents in colon cancer. The human B4GALNT2 gene drives the expression of a long and a short protein isoform (LF-B4GALNT2 and SF-B4GALNT2) sharing identical transmembrane and luminal domains. Both isoforms are trans-Golgi proteins and the LF-B4GALNT2 also localizes to post-Golgi vesicles thanks to its extended cytoplasmic tail. Control mechanisms underpinning Sd(a) and B4GALNT2 expression in the gastrointestinal tract are complex and not fully understood. This study reveals the existence of two unusual N-glycosylation sites in B4GALNT2 luminal domain. The first atypical N-X-C site is evolutionarily conserved and occupied by a complex-type N-glycan. We explored the influence of this N-glycan using site-directed mutagenesis and showed that each mutant had a slightly decreased expression level, impaired stability, and reduced enzyme activity. Furthermore, we observed that the mutant SF-B4GALNT2 was partially mislocalized in the endoplasmic reticulum, whereas the mutant LF-B4GALNT2 was still localized in the Golgi and post-Golgi vesicles. Lastly, we showed that the formation of homodimers was drastically impaired in the two mutated isoforms. An AlphaFold2 model of the LF-B4GALNT2 dimer with an N-glycan on each monomer corroborated these findings and suggested that N-glycosylation of each B4GALNT2 isoform controlled their biological activity.
format Online
Article
Text
id pubmed-9959560
institution National Center for Biotechnology Information
language English
publishDate 2023
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-99595602023-02-26 N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2 Cogez, Virginie Vicogne, Dorothée Schulz, Céline Portier, Lucie Venturi, Giulia de Ruyck, Jérôme Decloquement, Mathieu Lensink, Marc F. Brysbaert, Guillaume Dall’Olio, Fabio Groux-Degroote, Sophie Harduin-Lepers, Anne Int J Mol Sci Article The Sd(a) carbohydrate epitope and its biosynthetic B4GALNT2 enzyme are expressed in the healthy colon and down-regulated to variable extents in colon cancer. The human B4GALNT2 gene drives the expression of a long and a short protein isoform (LF-B4GALNT2 and SF-B4GALNT2) sharing identical transmembrane and luminal domains. Both isoforms are trans-Golgi proteins and the LF-B4GALNT2 also localizes to post-Golgi vesicles thanks to its extended cytoplasmic tail. Control mechanisms underpinning Sd(a) and B4GALNT2 expression in the gastrointestinal tract are complex and not fully understood. This study reveals the existence of two unusual N-glycosylation sites in B4GALNT2 luminal domain. The first atypical N-X-C site is evolutionarily conserved and occupied by a complex-type N-glycan. We explored the influence of this N-glycan using site-directed mutagenesis and showed that each mutant had a slightly decreased expression level, impaired stability, and reduced enzyme activity. Furthermore, we observed that the mutant SF-B4GALNT2 was partially mislocalized in the endoplasmic reticulum, whereas the mutant LF-B4GALNT2 was still localized in the Golgi and post-Golgi vesicles. Lastly, we showed that the formation of homodimers was drastically impaired in the two mutated isoforms. An AlphaFold2 model of the LF-B4GALNT2 dimer with an N-glycan on each monomer corroborated these findings and suggested that N-glycosylation of each B4GALNT2 isoform controlled their biological activity. MDPI 2023-02-18 /pmc/articles/PMC9959560/ /pubmed/36835549 http://dx.doi.org/10.3390/ijms24044139 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Cogez, Virginie
Vicogne, Dorothée
Schulz, Céline
Portier, Lucie
Venturi, Giulia
de Ruyck, Jérôme
Decloquement, Mathieu
Lensink, Marc F.
Brysbaert, Guillaume
Dall’Olio, Fabio
Groux-Degroote, Sophie
Harduin-Lepers, Anne
N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2
title N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2
title_full N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2
title_fullStr N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2
title_full_unstemmed N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2
title_short N-Glycan on the Non-Consensus N-X-C Glycosylation Site Impacts Activity, Stability, and Localization of the Sd(a) Synthase B4GALNT2
title_sort n-glycan on the non-consensus n-x-c glycosylation site impacts activity, stability, and localization of the sd(a) synthase b4galnt2
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9959560/
https://www.ncbi.nlm.nih.gov/pubmed/36835549
http://dx.doi.org/10.3390/ijms24044139
work_keys_str_mv AT cogezvirginie nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT vicognedorothee nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT schulzceline nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT portierlucie nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT venturigiulia nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT deruyckjerome nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT decloquementmathieu nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT lensinkmarcf nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT brysbaertguillaume nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT dalloliofabio nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT grouxdegrootesophie nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2
AT harduinlepersanne nglycanonthenonconsensusnxcglycosylationsiteimpactsactivitystabilityandlocalizationofthesdasynthaseb4galnt2