Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation
Glycosylation plays a critical role in various biological processes, yet identifying specific glycosyltransferase substrates remains a challenge due to the complexity of glycosylation. Here, we employ proximity labeling with biotin ligases BASU and TurboID to map the proximitome of MGAT3, a glycosyl...
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2025-01-01
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| author | Bowen Wang Xin He Yue Zhou Zengqi Tan Xiang Li Feng Guan Lei Lei |
| author_facet | Bowen Wang Xin He Yue Zhou Zengqi Tan Xiang Li Feng Guan Lei Lei |
| author_sort | Bowen Wang |
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| description | Glycosylation plays a critical role in various biological processes, yet identifying specific glycosyltransferase substrates remains a challenge due to the complexity of glycosylation. Here, we employ proximity labeling with biotin ligases BASU and TurboID to map the proximitome of MGAT3, a glycosyltransferase responsible for the biosynthesis of the bisecting GlcNAc structure, in HEK293T cells. This approach enriched 116 and 189 proteins, respectively, identifying 17 common substrates shared with bisecting GlcNAc-bearing proteome obtained via intact glycopeptide enrichment methods. Gene ontology analysis revealed that the enriched proteins were predominantly localized in the exosome, endoplasmic reticulum, and Golgi apparatus, consistent with subcellular localization of MGAT3 substrates. Notably, four novel substrates, GOLM2, CCDC134, ASPH, and ERO1A, were confirmed to bear bisecting GlcNAc modification, validating the utility of the proximity labeling method. Furthermore, we observed that bisecting GlcNAc modification inhibits breast cancer progression by promoting the degradation of α-galactosidase A (GLA). These findings demonstrate the efficacy of proximity labeling in identifying glycosyltransferase substrates and provide insights into the functional impact of bisecting GlcNAc modification. |
| format | Article |
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| institution | Kabale University |
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| language | English |
| publishDate | 2025-01-01 |
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| spelling | doaj-art-80135ffcc82c4e368aaa80d780e5de7c2025-01-24T13:26:41ZengMDPI AGCells2073-44092025-01-0114210310.3390/cells14020103Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA DegradationBowen Wang0Xin He1Yue Zhou2Zengqi Tan3Xiang Li4Feng Guan5Lei Lei6Key Laboratory of Resource Biology and Biotechnology Western China, Ministry of Education, Provincial Key Laboratory of Biotechnology, College of Life Sciences, Northwest University, Xi’an 710069, ChinaDepartment of Functional Laboratory, College of Laboratory Medicine, Dalian Medical University, Dalian 116044, ChinaKey Laboratory of Resource Biology and Biotechnology Western China, Ministry of Education, Provincial Key Laboratory of Biotechnology, College of Life Sciences, Northwest University, Xi’an 710069, ChinaInstitute of Hematology, School of Medicine, Northwest University, Xi’an 710069, ChinaInstitute of Hematology, School of Medicine, Northwest University, Xi’an 710069, ChinaKey Laboratory of Resource Biology and Biotechnology Western China, Ministry of Education, Provincial Key Laboratory of Biotechnology, College of Life Sciences, Northwest University, Xi’an 710069, ChinaKey Laboratory of Resource Biology and Biotechnology Western China, Ministry of Education, Provincial Key Laboratory of Biotechnology, College of Life Sciences, Northwest University, Xi’an 710069, ChinaGlycosylation plays a critical role in various biological processes, yet identifying specific glycosyltransferase substrates remains a challenge due to the complexity of glycosylation. Here, we employ proximity labeling with biotin ligases BASU and TurboID to map the proximitome of MGAT3, a glycosyltransferase responsible for the biosynthesis of the bisecting GlcNAc structure, in HEK293T cells. This approach enriched 116 and 189 proteins, respectively, identifying 17 common substrates shared with bisecting GlcNAc-bearing proteome obtained via intact glycopeptide enrichment methods. Gene ontology analysis revealed that the enriched proteins were predominantly localized in the exosome, endoplasmic reticulum, and Golgi apparatus, consistent with subcellular localization of MGAT3 substrates. Notably, four novel substrates, GOLM2, CCDC134, ASPH, and ERO1A, were confirmed to bear bisecting GlcNAc modification, validating the utility of the proximity labeling method. Furthermore, we observed that bisecting GlcNAc modification inhibits breast cancer progression by promoting the degradation of α-galactosidase A (GLA). These findings demonstrate the efficacy of proximity labeling in identifying glycosyltransferase substrates and provide insights into the functional impact of bisecting GlcNAc modification.https://www.mdpi.com/2073-4409/14/2/103bisecting GlcNAcMGAT3 substratesproximity labelingGLA |
| spellingShingle | Bowen Wang Xin He Yue Zhou Zengqi Tan Xiang Li Feng Guan Lei Lei Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation Cells bisecting GlcNAc MGAT3 substrates proximity labeling GLA |
| title | Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation |
| title_full | Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation |
| title_fullStr | Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation |
| title_full_unstemmed | Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation |
| title_short | Proximity Labeling-Based Identification of MGAT3 Substrates and Revelation of the Tumor-Suppressive Role of Bisecting GlcNAc in Breast Cancer via GLA Degradation |
| title_sort | proximity labeling based identification of mgat3 substrates and revelation of the tumor suppressive role of bisecting glcnac in breast cancer via gla degradation |
| topic | bisecting GlcNAc MGAT3 substrates proximity labeling GLA |
| url | https://www.mdpi.com/2073-4409/14/2/103 |
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