The Breast Cancer-Associated Glycoforms of MUC1, MUC1-Tn and sialyl-Tn, Are Expressed in COSMC Wild-Type Cells and Bind the C-Type Lectin MGL.

Richard Beatson,Gjertrud Maurstad,Appitha Arulappu,Julia Coleman,Henrik Clausen,Ulla Mandel,Gianfranco Picco,Hans H Wandell,Joy M Burchell,Joyce Taylor-Papadimitriou,Marit Sletmoen

doi:10.1371/journal.pone.0125994

Abstract

Aberrant glycosylation occurs in the majority of human cancers and changes in mucin-type O-glycosylation are key events that play a role in the induction of invasion and metastases. These changes generate novel cancer-specific glyco-antigens that can interact with cells of the immune system through carbohydrate binding lectins. Two glyco-epitopes that are found expressed by many carcinomas are Tn (GalNAc-Ser/Thr) and STn (NeuAcα2,6GalNAc-Ser/Thr). These glycans can be carried on many mucin-type glycoproteins including MUC1. We show that the majority of breast cancers carry Tn within the same cell and in close proximity to extended glycan T (Galβ1,3GalNAc) the addition of Gal to the GalNAc being catalysed by the T synthase. The presence of active T synthase suggests that loss of the private chaperone for T synthase, COSMC, does not explain the expression of Tn and STn in breast cancer cells. We show that MUC1 carrying both Tn or STn can bind to the C-type lectin MGL and using atomic force microscopy show that they bind to MGL with a similar deadadhesion force. Tumour associated STn is associated with poor prognosis and resistance to chemotherapy in breast carcinomas, inhibition of DC maturation, DC apoptosis and inhibition of NK activity. As engagement of MGL in the absence of TLR triggering may lead to anergy, the binding of MUC1-STn to MGL may be in part responsible for some of the characteristics of STn expressing tumours.

Highlights

Glycosylation is one of the most widely found and complex post-translational modifications, and the glycome encompasses a vast and extensive repertoire of sugars covalently linked to PLOS ONE | DOI:10.1371/journal.pone.0125994 May 7, 2015Tn and STn Antigen on MUC1 in Breast Cancer, Interaction with MGLLondon and King’s College Hospital NHS Foundation Trust
To investigate the role of COSMC in Tn expression, 40 cases of primary breast cancer were stained with antibodies to MUC1 (HMFG2), MUC1-Tn/STn (5E5) and MUC1-T (1B9), see S1 Table for the results of the individual tumors
To determine if Tn and T were in close proximity, the proximity ligation assay was applied to sections of primary breast cancers using 5E5 coupled to biotin and 1B9 coupled to digoxigenin

Summary

Introduction

Glycosylation is one of the most widely found and complex post-translational modifications, and the glycome encompasses a vast and extensive repertoire of sugars covalently linked to PLOS ONE | DOI:10.1371/journal.pone.0125994 May 7, 2015. Mucin-type O-linked glycosylation of proteins is one of the most diverse forms of glycosylation because it involves 50–100 distinct genes, including up to 20 polypeptide GalNAc-transferases that control where the O-glycans are attached. Over-expression of the ST3Gal-I sialyltransferase results in the expression of ST [13] and the expression of STn is due to the turning on of the transcription of another sialyltransferase, ST6GalNAc-I [14] It is not clear why the Tn glycan is so widely expressed a number of mechanisms have been reported. These include mutations in the gene encoding COSMC resulting in loss of T synthase activity [15], hypermethylation of COSMC in pancreatic cancer [16] and relocation of polypeptide GalNAc transferases to the ER [17]. 20–25% of breast cancer are positive for STn and it is found on the cell surface [12], permitting interaction with MGL expressing cells

Materials and Methods

Results

Discussion