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Implication of N-glycolylneuraminic acid in regulation of cell adhesiveness of C2C12 myoblast cells during differentiation into myotube cells

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Abstract

A transition of sialic acid (Sia) species on GM3 ganglioside from N-acetylneuraminic acid (Neu5Ac) to N-glycolylneuraminic acid (Neu5Gc) takes place in mouse C2C12 myoblast cells during their differentiation into myotube cells. However, the meaning of this Sia transition remains unclear. This study thus aims to gain a functional insight into this phenomenon. The following lines of evidence show that the increased de novo synthesis of Neu5Gc residues in differentiating myoblast cells promotes adhesiveness of the cells, which is beneficial for promotion of differentiation. First, the Sia transition occurred even in the C2C12 cells cultured in serum-free medium, indicating that it happens through de novo synthesis of Neu5Gc. Second, GM3(Neu5Gc) was localized in myoblast cells, but not in myotube cells, and related to expression of the CMP-Neu5Ac hydroxylase (CMAH) gene. Notably, expression of CMAH precedes myotube formation not only in differentiating C2C12 cells, but also in mouse developing embryos. Since the myoblast cells were attached on the dish surface more strongly than the myotube cells, expression of GM3(Neu5Gc) may be related to the surface attachment of the myoblast cells. Third, exogenous Neu5Gc, but not Neu5Ac, promoted differentiation of C2C12 cells, thus increasing the number of cells committed to fuse with each other. Fourth, the CMAH-transfected C2C12 cells were attached on the gelatin-coated surface much more rapidly than the mock-cells, suggesting that the expression of CMAH promotes cell adhesiveness through the expression of Neu5Gc.

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Acknowledgements

I (KK) am personally grateful to Dr. Sen-itiro Hakomori for his direct and indirect encouragements on me especially early in my career. I still remember my feeling when I happened to encounter his paper (Eggens et al. J. Biol. Chem. 1989; 264:9476-84) in the airplane from Japan to the US for attending an international meeting. It was as if I was struck by lightning. This was an early stage-study of the carbohydrate-carbohydrate interaction in the biological context, and actually overturned the existing concept of biomolecules. Since that time, I have been inspired by his way of thinking in science from time to time. No doubt that his contributions to glycobiology, particularly glycosphingolipid biology, would never been forgotten forever.

We thank Dr. Jin-ichi Inokuchi for his encouragement to this study. This work was supported by the Nagoya University operational subsidies 2019-2020 (to KK), Grants-in-Aid for JSPS Fellow 17J05805 (to SoG) and for Scientific Research (C) 18K06120 (to SnG).

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Authors and Affiliations

  1. Graduate School of Bioagricultural Sciences and Bioscience and Biotechnology Center, Nagoya University, Nagoya, 464-8601, Japan

    Shiori Go, Chihiro Sato, Masaya Hane & Ken Kitajima

  2. Institute for Glyco-Core Research (iGCORE), Nagoya University, Nagoya, 464-8601, Japan

    Shiori Go, Chihiro Sato, Masaya Hane, Shinji Go & Ken Kitajima

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  1. Shiori Go
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Contributions

K.K. designed the study, analyzed the data, and prepared the main manuscript text and figures. So.G. was involved in all the experimental design and performances, and prepared the main manuscript text and all the figures. M.H. was involved in the design of construction of plasmids and management of the animal experiments; Sn.G. designed the study and analyzed the data. S.C. designed the study and analyzed the data.

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Correspondence to Ken Kitajima.

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Go, S., Sato, C., Hane, M. et al. Implication of N-glycolylneuraminic acid in regulation of cell adhesiveness of C2C12 myoblast cells during differentiation into myotube cells. Glycoconj J 39, 619–631 (2022). https://doi.org/10.1007/s10719-022-10049-9

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