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Functional Characterization of POFUT1 Variants Associated with Colorectal Cancer.


ABSTRACT: BACKGROUND:Protein O-fucosyltransferase 1 (POFUT1) overexpression, which is observed in many cancers such as colorectal cancer (CRC), leads to a NOTCH signaling dysregulation associated with the tumoral process. In rare CRC cases, with no POFUT1 overexpression, seven missense mutations were found in human POFUT1. METHODS:Recombinant secreted forms of human WT POFUT1 and its seven mutated counterparts were produced and purified. Their O-fucosyltransferase activities were assayed in vitro using a chemo-enzymatic approach with azido-labeled GDP-fucose as a donor substrate and NOTCH1 EGF-LD26, produced in E. coli periplasm, as a relevant acceptor substrate. Targeted mass spectrometry (MS) was carried out to quantify the O-fucosyltransferase ability of all POFUT1 proteins. FINDINGS:MS analyses showed a significantly higher O-fucosyltransferase activity of six POFUT1 variants (R43H, Y73C, T115A, I343V, D348N, and R364W) compared to WT POFUT1. INTERPRETATION:This study provides insights on the possible involvement of these seven missense mutations in colorectal tumors. The hyperactive forms could lead to an increased O-fucosylation of POFUT1 protein targets such as NOTCH receptors in CRC patients, thereby leading to a NOTCH signaling dysregulation. It is the first demonstration of gain-of-function mutations for this crucial glycosyltransferase, modulating NOTCH activity, as well as that of other potential glycoproteins.

SUBMITTER: Deschuyter M 

PROVIDER: S-EPMC7352195 | biostudies-literature | 2020 May

REPOSITORIES: biostudies-literature

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Functional Characterization of POFUT1 Variants Associated with Colorectal Cancer.

Deschuyter Marlène M   Pennarubia Florian F   Pinault Emilie E   Legardinier Sébastien S   Maftah Abderrahman A  

Cancers 20200531 6


<h4>Background</h4>Protein <i>O</i>-fucosyltransferase 1 (POFUT1) overexpression, which is observed in many cancers such as colorectal cancer (CRC), leads to a NOTCH signaling dysregulation associated with the tumoral process. In rare CRC cases, with no <i>POFUT1</i> overexpression, seven missense mutations were found in human POFUT1.<h4>Methods</h4>Recombinant secreted forms of human WT POFUT1 and its seven mutated counterparts were produced and purified. Their <i>O</i>-fucosyltransferase activ  ...[more]

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