FUT8-mediated aberrant N-glycosylation of SEMA7A promotes head and neck squamous cell carcinoma progression

Abstract

SEMA7A belongs to the Semaphorin family and is involved in the oncogenesis and tumor progression. Aberrant glycosylation has been intricately linked with immune escape and tumor growth. SEMA7A is a highly glycosylated protein with five glycosylated sites. The underlying mechanisms of SEMA7A glycosylation and its contribution to immunosuppression and tumorigenesis are unclear. Here, we identify overexpression and aberrant N-glycosylation of SEMA7A in head and neck squamous cell carcinoma, and elucidate fucosyltransferase FUT8 catalyzes aberrant core fucosylation in SEMA7A at N-linked oligosaccharides (Asn 105, 157, 258, 330, and 602) via a direct protein‒protein interaction. A glycosylated statue of SEMA7A is necessary for its intra-cellular trafficking from the cytoplasm to the cytomembrane. Cytokine EGF triggers SEMA7A N-glycosylation through increasing the binding affinity of SEMA7A toward FUT8, whereas TGF-β1 promotes abnormal glycosylation of SEMA7A via induction of epithelial-mesenchymal transition. Aberrant N-glycosylation of SEMA7A leads to the differentiation of CD8⁺ T cells along a trajectory toward an exhausted state, thus shaping an immunosuppressive microenvironment and being resistant immunogenic cell death. Deglycosylation of SEMA7A significantly improves the clinical outcome of EGFR-targeted and anti-PD-L1-based immunotherapy. Finally, we also define RBM4, a splice regulator, as a downstream effector of glycosylated SEMA7A and a pivotal mediator of PD-L1 alternative splicing. These findings suggest that targeting FUT8-SEMA7A axis might be a promising strategy for improving antitumor responses in head and neck squamous cell carcinoma patients.

References

1. Siegel R. L., Miller K. D., Fuchs H. E.& Jemal, A. Cancer statistics, 2022.CA Cancer J. Clin. 72, 7-33 (2022).
2. Magalhaes, A., Duarte, H. O.& Reis, C. A. Aberrant glycosylation in cancer: a novel molecular mechanism controlling metastasis.Cancer Cell 31, 733-735 (2017).
3. RodrÍguez, E., Schetters, S. T.T. & van Kooyk, Y. The tumour glyco-code as a novel immune checkpoint for immunotherapy.Nat. Rev. Immunol. 18, 204-211 (2018).
4. Oliveira-Ferrer, L., Legler, K. & Milde-Langosch, K. Role of protein glycosylation in cancer metastasis.Semin. Cancer Biol. 44, 141-152 (2017).
5. Marsico G., Russo L., Quondamatteo F.& Pandit, A. Glycosylation and integrin regulation in cancer.Trends Cancer 4, 537-552 (2018).
6. Chakraborty A.& Dimitroff, C. J. Cancer immunotherapy needs to learn how to stick to its guns.J. Clin. Invest. 129, 5089-5091 (2019).
7. Gomes J., Stefan M., Ana M.& Celso, A. R. Early GalNAc O-glycosylation: pushing the tumor boundaries.Cancer Cell 32, 544-545 (2017).
8. Li, C. W.et al.Glycosylation and stabilization of programmed death ligand-1 suppresses T-cell activity.Nat. Commun. 7, 12632(2016).
9. Sasawatari S., Okamoto Y., Kumanogoh A.& Toyofuku, T. Blockade of N-glycosylation promotes antitumor immune response of T cells.J. Immunol. 204, 1373-1385 (2020).
10. Shao, B.et al.Deglycosylation of PD-L1 by 2-deoxyglucose reverses PARP inhibitor-induced immunosuppression in triple-negative breast cancer.Am. J. Cancer. Res. 8, 1837-1846 (2018).
11. Li, C. W.et al. Eradication of triple-negative breast cancer cells by targeting glycosylated PD-L1. Cancer Cell 33, 187-201.e10 (2018).
12. Lee, H. H.et al. Removal of N-linked glycosylation enhances PD-L1 detection and predicts Anti-PD-1/PD-L1 therapeutic efficacy. Cancer Cell 36, 168-178.e4 (2019).
13. Sun, L.et al.Targeting Glycosylated PD-1 induces potent antitumor immunity.Cancer Res. 80, 2298-2310 (2020).
14. Chen, J. T.et al.Glycoprotein B7-H3 overexpression and aberrant glycosylation in oral cancer and immune response.Proc. Natl Acad. Sci. USA 112, 13057-13062 (2015).
15. Huang, Y.et al.FUT8-mediated aberrant N-glycosylation of B7H3 suppresses the immune response in triple-negative breast cancer.Nat. Commun. 12, 2672(2021).
16. Xu, Y.et al.PD-L2 glycosylation promotes immune evasion and predicts anti-EGFR efficacy.J. Immunother. Cancer 9, e002699(2021).
17. Lin M. C., Huang M. J., Liu C. H., Yang T. L.& Huang, M. C. GALNT2 enhances migration and invasion of oral squamous cell carcinoma by regulating EGFR glycosylation and activity.Oral Oncol. 50, 478-484 (2014).
18. Song, X.et al.Pharmacologic suppression of B7-H4 glycosylation restores antitumor immunity in immune-cold breast cancers.Cancer Discov. 10, 1872-1893 (2020).
19. Jongbloets, B. C.et al.Stage-specific functions of Semaphorin7A during adult hippocampal neurogenesis rely on distinct receptors.Nat. Commun. 8, 14666(2017).
20. van Rijn, A.et al. Semaphorin 7A promotes chemokine-driven dendritic cell migration.J. Immunol. 196, 459-468 (2016).
21. Yamada, A.et al.Molecular cloning of a glycosylphosphatidylinositol-anchored molecule CD108w.J. Immunol. 162, 4094-4100 (1999).
22. Liu, H.et al.Structural basis of semaphorin-plexin recognition and viral mimicry from SEMA7A and A39R complexes with PlexinC1.Cell 142, 749-761 (2010).
23. Suzuki, K.et al.Semaphorin 7A initiates T-cell-mediated inflammatory responses through alpha1beta1 integrin.Nature 446, 680-684 (2007).
24. Suzuki, K., Kumanogoh, A.& Kikutani, H. Semaphorins and their receptors in immune cell interactions.Nat. Immunol. 9, 17-23 (2008).
25. Elder, A. M.et al.Semaphorin 7A promotes macrophage-mediated lymphatic remodeling during postpartum mammary gland involution and in breast cancer.Cancer Res. 78, 6473-6485 (2018).
26. Garcia-Areas, R. et al. Suppression of tumor-derived Semaphorin 7A and genetic ablation of host-derived Semaphorin 7A impairs tumor progression in a murine model of advanced breast carcinoma.Int. J. Oncol. 51, 1395-1404 (2017).
27. Garcia-Areas, R. et al. Semaphorin7A promotes tumor growth and exerts a pro-angiogenic effect in macrophages of mammary tumor-bearing mice.Front. Physiol. 5, 17(2014).
28. Tarullo, S. E.et al.Postpartum breast cancer progression is driven by semaphorin 7a-mediated invasion and survival.Oncogene 39, 2772-2785 (2020).
29. Kinehara, Y.et al.Semaphorin 7A promotes EGFR-TKI resistance in EGFR mutant lung adenocarcinoma cells.JCI Insight 3, e123093(2018).
30. Inoue N., Nishizumi H., Naritsuka H., Kiyonari H.& Sakano, H. SEMA7A/PlxnCl signaling triggers activity-dependent olfactory synapse formation.Nat. Commun. 9, 1842(2018).
31. Janssen, B. J.et al.Structural basis of semaphorin-plexin signalling.Nature 467, 1118-1122 (2010).
32. Grigaravicius P., von Deimling, A. & Frappart, P. O. RINT1 functions as a multitasking protein at the crossroads between genomic stability, ER homeostasis, and autophagy.Autophagy 12, 1413-1415 (2016).
33. Ng, B. G.et al.Expanding the molecular and clinical phenotypes of FUT8-CDG.J. Inherit. Metab. Dis. 43, 871-879 (2020).
34. Järvå, M. A.et al.Structural basis of substrate recognition and catalysis by fucosyltransferase 8.J. Biol. Chem. 295, 6677-6688 (2020).
35. Lin J. C.& Tarn, W. Y. RBM4 down-regulates PTB and antagonizes its activity in muscle cell-specific alternative splicing.J. Cell Biol. 193, 509-520 (2011).
36. D, D., Hung, K. Y. & Tarn, W. Y. RBM4 modulates radial migration via alternative splicing of Dab1 during cortex development.Mol. Cell Biol. 38, e00007-e00018 (2018).
37. Markus, M. A., Yang, Y. H.& Morris, B. J. Transcriptome-wide targets of alternative splicing by RBM4 and possible role in cancer.Genomics 107, 138-144 (2016).
38. Mahoney, K. M.et al.A secreted PD-L1 splice variant that covalently dimerizes and mediates immunosuppression.Cancer Immunol. Immunother. 68, 421-432 (2019).
39. Hassounah, N. B.et al.Identification and characterization of an alternative cancer-derived PD-L1 splice variant.Cancer Immunol. Immunother. 68, 407-420 (2019).
40. Wang, C.et al.Distinct roles of programmed death ligand 1 alternative splicing isoforms in colorectal cancer.Cancer Sci. 112, 178-193 (2021).
41. Cagnoni A. J.,Pérez Sáez, J. M., Rabinovich, G. A. & Mariño, K. V. Turning-off signaling by siglecs, selectins, and galectins: chemical inhibition of glycan-dependent interactions in cancer.Front Oncol 6, 109(2016).
42. Chen, C. Y.et al.Fucosyltransferase 8 as a functional regulator of nonsmall cell lung cancer.Proc. Natl Acad. Sci. USA 110, 630-635 (2013).
43. Agrawal, P.et al. A systems biology approach identifies FUT8 as a driver of melanoma metastasis. Cancer Cell 31, 804-819.e7 (2017).
44. Chiang, W. F.et al.Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) promotes EGF receptor signaling of oral squamous cell carcinoma metastasis via the complex N-glycosylation.Oncogene 37, 116-127 (2018).