Prof. Adnan Halim - Protein O-mannosylation: structure, function and cancer
Dr. Adnan Halim - Protein O-mannosylation: structure, function and cancer
Adnan Halim
Department of Cellular and Molecular Medicine, University of Copenhagen, DK
Protein O-mannosylation: structure, function and cancer
Protein O linked mannose (O Man) glycosylation is essential for mammalian development, and disruption of its biosynthesis causes severe muscular, neurological, and cardiac disease. O Man initiation is mediated by three distinct enzyme systems in humans—POMT1/POMT2, TMTC1–4, and TMEM260—which selectively modify specific classes of transmembrane proteins, including α dystroglycan, cadherins, plexins, and receptor tyrosine kinases. However, the principles governing substrate selection and pathway specificity have remained poorly defined.
By integrating quantitative O-glycoproteomics, genetic dissection, and structural analysis, we establish a framework for understanding mammalian O Man initiation and functions. Targeted glycoproteomics in glycoengineered human cells reveal previously unrecognized substrates of the POMT1/POMT2 complex and demonstrate that TMTC1–4 and TMEM260 preferentially modify Ig like domains of plasma membrane proteins involved in cell–cell and cell–matrix communication, supporting the emerging concept of domain restricted O Man glycosylation. To elucidate the molecular basis of this specificity, we determined the CryoEM structure of human TMEM260 in substrate bound states. Structures of TMEM260 complexed with its native donor dolichyl phosphate β-mannose and an acceptor peptide from plexin B2 reveal the architecture of a mammalian O-mannosyltransferase captured during catalysis. We identify a conserved O-Man acceptor sequon and demonstrate that TMEM260 preferentially modifies extended polypeptide segments, consistent with co-translational glycosylation.
Extending our studies to cancer, we show that O-Man glycosylated fusion partners play an active role in oncogenesis. In the KIAA1549::BRAF fusion, the most common driver in pediatric brain tumors, the KIAA1549 partner confers a specific dependency on POMT1/2. Genetic or pharmacologic inhibition of POMT1/2, responsible for O-Man glycosylation of the KIAA1549 partner, blocks fusion driven transformation by preventing glycosylation and maturation of the fusion protein, revealing a MAPK independent therapeutic vulnerability and highlighting fusion partners as actionable targets in cancer.
This seminar will thus highlight insight into CryoEM and structural mechanisms for O-Man biosynthesis, cellular functions, glycoengineering, and analytic platforms that have recently uncovered unexpected roles for O-Man in development and cancer..
References
- Cifuente et al. Structure and mechanism of the human TMEM260 O-mannosyltransferase. biorXiv (2026)
- Misek et al. O-mannosylation and protein maturation check-points represent therapeutic opportunities in BRAF fusion protein oncogenesis. biorXiv (2025)
- Povoolo et al. Global View of Domain-Specific O-Linked Mannose Glycosylation in Glycoengineered Cells. (2024) Mol Cell Proteomics
- Larsen et al. The SHDRA syndrome-associated gene TMEM260 encodes a protein-specific O-mannosyltransferase. (2023) PNAS
- Larsen et al. Discovery of an O-mannosylation pathway selectively serving cadherins and protocadherins. (2017) PNAS
Prof. Adnan Halim - Protein O-mannosylation: structure, function and cancer
Dr. Adnan Halim - Protein O-mannosylation: structure, function and cancer
Adnan Halim
Department of Cellular and Molecular Medicine, University of Copenhagen, DK
Protein O-mannosylation: structure, function and cancer
Protein O linked mannose (O Man) glycosylation is essential for mammalian development, and disruption of its biosynthesis causes severe muscular, neurological, and cardiac disease. O Man initiation is mediated by three distinct enzyme systems in humans—POMT1/POMT2, TMTC1–4, and TMEM260—which selectively modify specific classes of transmembrane proteins, including α dystroglycan, cadherins, plexins, and receptor tyrosine kinases. However, the principles governing substrate selection and pathway specificity have remained poorly defined.
By integrating quantitative O-glycoproteomics, genetic dissection, and structural analysis, we establish a framework for understanding mammalian O Man initiation and functions. Targeted glycoproteomics in glycoengineered human cells reveal previously unrecognized substrates of the POMT1/POMT2 complex and demonstrate that TMTC1–4 and TMEM260 preferentially modify Ig like domains of plasma membrane proteins involved in cell–cell and cell–matrix communication, supporting the emerging concept of domain restricted O Man glycosylation. To elucidate the molecular basis of this specificity, we determined the CryoEM structure of human TMEM260 in substrate bound states. Structures of TMEM260 complexed with its native donor dolichyl phosphate β-mannose and an acceptor peptide from plexin B2 reveal the architecture of a mammalian O-mannosyltransferase captured during catalysis. We identify a conserved O-Man acceptor sequon and demonstrate that TMEM260 preferentially modifies extended polypeptide segments, consistent with co-translational glycosylation.
Extending our studies to cancer, we show that O-Man glycosylated fusion partners play an active role in oncogenesis. In the KIAA1549::BRAF fusion, the most common driver in pediatric brain tumors, the KIAA1549 partner confers a specific dependency on POMT1/2. Genetic or pharmacologic inhibition of POMT1/2, responsible for O-Man glycosylation of the KIAA1549 partner, blocks fusion driven transformation by preventing glycosylation and maturation of the fusion protein, revealing a MAPK independent therapeutic vulnerability and highlighting fusion partners as actionable targets in cancer.
This seminar will thus highlight insight into CryoEM and structural mechanisms for O-Man biosynthesis, cellular functions, glycoengineering, and analytic platforms that have recently uncovered unexpected roles for O-Man in development and cancer..
References
- Cifuente et al. Structure and mechanism of the human TMEM260 O-mannosyltransferase. biorXiv (2026)
- Misek et al. O-mannosylation and protein maturation check-points represent therapeutic opportunities in BRAF fusion protein oncogenesis. biorXiv (2025)
- Povoolo et al. Global View of Domain-Specific O-Linked Mannose Glycosylation in Glycoengineered Cells. (2024) Mol Cell Proteomics
- Larsen et al. The SHDRA syndrome-associated gene TMEM260 encodes a protein-specific O-mannosyltransferase. (2023) PNAS
- Larsen et al. Discovery of an O-mannosylation pathway selectively serving cadherins and protocadherins. (2017) PNAS
