Dépôt numérique

AP‐1 mediated trafficking of the lysosomal vitamin B12 transporter ABCD4.

Sauvageau, Etienne; Castonguay, Annie ORCID logoORCID: https://orcid.org/0000-0001-5705-6353 et Lefrancois, Stephane . AP‐1 mediated trafficking of the lysosomal vitamin B12 transporter ABCD4. In: ASCB/EMBO Meeting, 2-6 décembre 2017, Philadelphie.

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A variety of human diseases are caused by improper protein trafficking within the trans‐Golgi network(TGN)‐endosome‐ lysosome system. Several bacterial toxins and viruses also employ this transport system in order to infect cells. Fundamental understanding of the mechanisms regulating this trafficking pathway is therefore essential to the development of novel therapeutic approaches. MEDNIK syndrome, a rare genetic disease characterized by mental retardation, enteropathy, deafness, peripheral neuropathy, ichthyosis and keratodermia, is caused by mutations in the AP1S1 gene encoding for the σ1A subunit of the clathrin adaptor complex AP‐1. This heterotetrameric clathrin adaptor complex is involved in recruiting cargo proteins and initiating the formation of transport vesicles between the TGN and endosomes‐lysosomes. Cytosolic AP‐1 is recruited by the small GTPase Arf1 to the membranes of the TGN where it interacts with specific motifs within the cytosolic tail of transmembrane proteins while also recruiting clathrin and inducing the formation of clathrin‐coated vesicles(CCVs). Although defective trafficking of the ATP7A copper pump and the copper‐dependent enzyme PAM have been shown to contribute to the pathogenic mechanism of MEDNIK syndrome, the severity and complexity of the disease, coupled with the broad function of AP‐1, suggest that aberrant transport of other proteins also contribute to the syndrome. Mutations in the lysosomal transmembrane protein ABCD4 block vitamin B12 transport to the cytosol and cause a disease sharing clinical manifestations (neuropathy, hypotonia) with MEDNIK syndrome. Using co‐immunoprecipitation and bioluminescence resonance energy transfer(BRET), we found that ABCD4 interacts withAP‐1 in living cells. Mutations of the classical tyrosine(YXXØ) or dihydrophobically ([DE]XXXL[LI]) based motifs in the c‐terminal tail of ABCD4 did not lead to a loss of interaction with AP‐1, suggesting a novel mode of binding. BRET experiments in an Arf1‐KO CRISPR generated HEK293T cell line display reduced levels of interaction between ABCD4 and the μ1 and β1 subunits of AP‐1. Moreover, ABCD4 accumulates in the Golgi apparatus in a γ‐KOCRISPR generated cell line as determined by fluorescence microscopy. Finally,fluorescence microscopy experiments using bodipy‐conjugated vitamin B12 show lysosomal accumulation of vitamin B12 in fibroblasts isolated from a MEDNIK patient. Together, these results suggest that ABCD4 transport to the lysosomes requires AP‐1 and that aberrant vitamin B12 metabolism could contribute to MEDNIK syndrome pathogenicity.

Type de document: Document issu d'une conférence ou d'un atelier
Informations complémentaires: Poster, P3132
Mots-clés libres: -
Centre: Centre INRS-Institut Armand Frappier
Date de dépôt: 16 août 2018 02:59
Dernière modification: 21 févr. 2022 17:11
URI: https://espace.inrs.ca/id/eprint/7495

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