Abstract
Glycosylation, the most prevalent and diverse post-translational modification of protein, plays crucial biological roles in many physiological and pathological events. Alteration of N-glycan has been detected during breast cancer progression. Among the specific N-glycan structures, bisecting N-Acetylglucosamine (GlcNAc) is a β1,4-linked GlcNAc attached to the core β-mannose residue, and is catalyzed by glycosyltransferase MGAT3. Bisecting GlcNAc levels were commonly dysregulated in different types of cancer. In this study, we utilized mass spectrometry and lectin microarray analysis to investigate aberrant N-glycans in breast cancer cells. Our data showed the decreased levels of bisecting GlcNAc and down-regulated expression of MGAT3 in breast cancer cells than normal epithelial cells. Using PHA-E (a plant lectin recognizing and combining bisecting GlcNAc) based enrichment coupled with nanoLC-MS/MS, we analyzed the glycoproteins bearing bisecting GlcNAc in various breast cancer cells. Among the differentially expressed glycoproteins, levels of bisecting GlcNAc on EGFR were significantly decreased in breast cancer cells, confirmed by immunostaining and immunoprecipitation. We overexpressed MGAT3 in breast cancer MDA-MB-231 cells, and overexpression of MGAT3 significantly enhanced the bisecting N-GlcNAc on EGFR and suppressed the EGFR/Erk signaling, which further resulted in the reduction of migratory ability, cell proliferation, and clonal formation. Taken together, we conclude that bisecting N-GlcNAc on EGFR inhibits malignant phenotype of breast cancer via down-regulation of EGFR/Erk signaling.
Highlights
Breast cancer (BCa) is one of the most common cancers and associated with high mortality rates in women [1, 2]
The results showed that bisecting GlcNAc could retard the cancerous phenotype of BCa cells by suppressing EGFR/ERK signaling
Aberrant glycosylation is an established hallmark of BCa such as CA 16-3, carcinoembryonic antigen (CEA)
Summary
Breast cancer (BCa) is one of the most common cancers and associated with high mortality rates in women [1, 2]. Blood biomarkers of BCa for early detection is widely studied for improvement of prognosis and survival rate. Some blood-borne tumor markers were widely used for screening, Bisecting GlcNAc Inhibits Malignant Phenotype monitoring, and prognosis of BCa patients, for example, carbohydrate antigen 15-3 (CA15-3) [4, 5] and carcinoembryonic antigen (CEA) [6]. We previously observed decreased levels of bisecting N-acetylglucosamine (GlcNAc) and its glycosyltransferase N-acetylglucosaminyltransferase III (GlcNAcT- III, termed as MGAT3) expression in TGFβ- and hypoxia- induced EMT process [24, 25]. We investigated the levels of bisecting GlcNAc in various breast cells using high-throughput techniques (MALDI-TOF/TOF-MS and lectin microarray), identified the target proteins bearing bisecting GlcNAc using lectin PHA-E enrichment coupled with NanoLC-MS/MS analysis, and explored the effects of bisecting GlcNAc on target protein
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