The Mechanism of Stimulation of Myogenesis under the Action of Succinic Acid Through the Succinate Receptor SUCNR1

Cover Page

Cite item

Full Text

Open Access Open Access
Restricted Access Access granted
Restricted Access Subscription Access

Abstract

In a study on cells of the C2C12 line, the effect of succinic acid on the processes of myogenesis was studied. In the concentration range of 10-1000 microns, succinic acid stimulated the process of myogenic differentiation, increasing the number of myogenesis factors MyoD (at all stages of myogenesis) and myogenin (at the stage of terminal differentiation). The Western blot method revealed specific succinate receptors SUCNR1 in C2C12 cells, the level of which decreased during myogenesis. When succinic acid was added to cells, the level of intracellular succinate did not change significantly and decreased during myogenic differentiation. Using a specific Gai protein inhibitor, pertussis toxin, it was found that stimulation of myogenesis of C2C12 under the action of succinic acid is realized through SUCNR1–Gai.

Full Text

Restricted Access

About the authors

Yu. V. Abalenikhina

Ryazan State Medical University

Author for correspondence.
Email: abalenihina88@mail.ru
Russian Federation, Ryazan

M. O. Isaeva

Ryazan State Medical University

Email: abalenihina88@mail.ru
Russian Federation, Ryazan

P. Yu. Mylnikov

Ryazan State Medical University

Email: abalenihina88@mail.ru
Russian Federation, Ryazan

A. V. Shchulkin

Ryazan State Medical University

Email: abalenihina88@mail.ru
Russian Federation, Ryazan

E. N. Yakusheva

Ryazan State Medical University

Email: abalenihina88@mail.ru
Russian Federation, Ryazan

References

  1. Xu, M., Chen, X., Chen, D., Yu, B., Li, M., He, J., and Huang, Z. (2020) Regulation of skeletal myogenesis by microRNAs, J. Cell. Physiol., 235, 87-104, https://doi.org/10.1002/jcp.28986.
  2. Arnold, H. H., and Braun, T. (1996) Targeted inactivation of myogenic factor genes reveals their role during mouse myogenesis: a review, Int. J. Dev. Biol., 40, 345-353.
  3. Lassar, A. B., Skapek, S. X., and Novitch, B. (1994) Regulatory mechanisms that coordinate skeletal muscle differentiation and cell cycle withdrawal, Curr. Opin. Cell Biol., 6, 788-794, https://doi.org/10.1016/0955-0674(94)90046-9.
  4. Cuenda, A., and Cohen, P. (1999) Stress-activated protein kinase-2/p38 and a rapamycin-sensitive pathway are required for C2C12 myogenesis, J. Biol. Chem., 274, 4341-4346, https://doi.org/10.1074/jbc.274.7.4341.
  5. Buckingham, M., and Vincent, S. D. (2009) Distinct and dynamic myogenic populations in the vertebrate embryo, Curr. Opin. Genet. Dev., 19, 444-453, https://doi.org/10.1016/j.gde.2009.08.001.
  6. Arneson-Wissink, P. C., Hogan, K. A., Ducharme, A. M., Samani, A., Jatoi, A., and Doles, J. D. (2020) The wasting-associated metabolite succinate disrupts myogenesis and impairs skeletal muscle regeneration, JCSM Rapid Commun., 3, 56-69, https://doi.org/10.1002/rco2.14.
  7. Fredriksson, R., Lagerström, M. C., Lundin, L.-G., and Schiöth, H. B. (2003) The G-protein-coupled receptors in the human genome form five main families. Phylogenetic analysis, paralogon groups, and fingerprints, Mol. Pharmacol., 63, 1256-1272, https://doi.org/10.1124/mol.63.6.1256.
  8. Wang, T., Xu, Y. Q., Yuan, Y. X., Xu, P. W., Zhang, C., Li, F., Wang, L. N., Yin, C., Zhang, L., Cai, X. C., Zhu, C. J., Xu, J. R., Liang, B. Q., Schaul, S., Xie, P. P., Yue, D., Liao, Z. R., Yu, L. L., Luo, L., Zhou, G., Yang, J. P., He, Z. H., Du, M., Zhou, Y. P., Deng, B. C., Wang, S. B., Gao, P., Zhu, X. T., Xi, Q. Y., et al. (2019) Succinate induces skeletal muscle fiber remodeling via SUCNR1 signaling, EMBO Rep., 9, e47892, https://doi.org/10.15252/embr.201947892.
  9. Abdelmoez, A. M., Dmytriyeva, O., Zurke, Y. X., Trauelsen, M., Marica, A. A., Savikj, M., Smith, J. A. B., Monaco, C., Schwartz, T. W., Krook, A., and Pillon, N. J. (2023) Cell selectivity in succinate receptor SUCNR1/GPR91 signaling in skeletal muscle, Am. J. Physiol. Endocrinol. Metab., 324, E289-E298, https://doi.org/10.1152/ajpendo.00009.2023.
  10. He, W., Miao, F. J., Lin, D. C., Schwandner, R. T., Wang, Z., Gao, J., Chen, J. L., Tian, H., and Ling, L. (2004) Citric acid cycle intermediates as ligands for orphan G-protein-coupled receptors, Nature, 429, 188-193, https://doi.org/10.1038/nature02488.
  11. Gilissen, J., Jouret, F., Pirotte, B., and Hanson, J. (2016) Insight into SUCNR1 (GPR91) structure and function, Pharmacol. Ther., 159, 56-65, https://doi.org/10.1016/j.pharmthera.2016.01.008.
  12. Harden T. K., Waldo G. L., Hicks S. N., and Sondek J. (2011) Mechanism of activation and inactivation of Gq/phospholipase C-β signaling nodes, Chem. Rev., 10, 6120-6129, https://doi.org/10.1021/cr200209p.
  13. Yaffe, D., and Saxel, O. (1977) A myogenic cell line with altered serum requirements for differentiation, Differentiation, 7, 159-166, https://doi.org/10.1111/j.1432-0436.1977.tb01507.x.
  14. Sin, J., Andres, A. M., Taylor, D. J., Weston, T., Hiraumi, Y., Stotland, A., Kim, B. J., Huang, C., Doran, K. S., and Gottlieb, R. A. (2016) Mitophagy is required for mitochondrial biogenesis and myogenic differentiation of C2C12 myoblasts, Autophagy, 12, 369-380, https://doi.org/10.1080/15548627.2015.1115172.
  15. Исаева М. О., Гаджиева Ф. Т., Абаленихина Ю. В., Щулькин А. В., Якушева Е. Н. (2023) Способ культивирования и механизмы регуляции этапов миогенеза клеточной линии С2С12, Российский медико-биологический вестник им. академика И.П. Павлова, 4, 525-534, https://doi.org/10.17816/PAVLOVJ375362.
  16. Буев Д. О., Емелин А. М., Яковлев И. А., Деев Р. В. (2020) Культивирование миобластов и миосателлитоцитов in vitro, Наука молодых (Eruditio Juvenium), 1, 86-97, https://doi.org/10.23888/HMJ20208186-97.
  17. Sundström, L., Greasley, P. J., Engberg, S., Wallander, M., and Ryberg, E. (2013) Succinate receptor GPR91, a Gα(i) coupled receptor that increases intracellular calcium concentrations through PLCβ, FEBS Lett., 15, 2399-2404, https://doi.org/10.1016/j.febslet.2013.05.067.
  18. Емелин А. М., Буев Д. О., Слабикова А. А., Яковлев И. А., Деев Р. В. (2019) Количественная оценка миогенной дифференцировки клеточной линии С2С12 с использованием полиэтиленгликоля и индуцированных сред in vitro, Гены Клетки, 14, 87.
  19. Sestili, P., Barbieri, E., Martinelli, C., Battistelli, M., Guescini, M., Vallorani, L., Casadei, L., D’Emilio, A., Falcieri, E., Piccoli, G., Agostini, D., Annibalini, G., Paolillo, M., Gioacchini, A. M., and Stocchi, V. (2009) Creatine supplementation prevents the inhibition of myogenic differentiation in oxidatively injured C2C12 murine myoblasts, Mol. Nutr. Food Res., 9, 1187-1204, https://doi.org/10.1002/mnfr.200800504.
  20. Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding, Anal. Biochem., 72, 248-254, https://doi.org/10.1006/abio.1976.9999.
  21. Kohout, T. A., and Lefkowitz, R. J. (2003) Regulation of G protein-coupled receptor kinases and arrestins during receptor desensitization, Mol. Pharmacol., 1, 9-18, https://doi.org/10.1124/mol.63.1.9.
  22. Vercellino, I., and Sazanov, L. A. (2022) The assembly, regulation and function of the mitochondrial respiratory chain, Nat. Rev. Mol. Cell Biol., 2, 141-161, https://doi.org/10.1038/s41580-021-00415-0.
  23. Locht, C., and Antoine, R. (1995) A proposed mechanism of ADP-ribosylation catalyzed by the pertussis toxin S1 subunit, Biochimie, 5, 333-540, https://doi.org/10.1016/0300-9084(96)88143-0.
  24. Najimi, M., Gailly, P., Maloteaux, J. M., and Hermans, E. (2002) Distinct regions of C-terminus of the high affinity neurotensin receptor mediate the functional coupling with pertussis toxin sensitive and insensitive G-proteins, FEBS Lett., 1-3, 329-333, https://doi.org/10.1016/s0014-5793(02)02285-8.
  25. Siow, N. L., Choi, R. C. Y., Cheng, A. W. M., Jiang, J. X. S., Wan, D. C. C., Zhu, S. Q., and Tsim, K. W. K. (2002) A Cyclic AMP-dependent pathway regulates the expression of acetylcholinesterase during myogenic differentiation of C2C12 cells, J. Biol. Chem., 39, 36129-36136, doi: 10.1074/jbc.M206498200.
  26. Kitzmann, M., Vandromme, M., Schaeffer, V., Carnac, G., Labbé, J. C., Lamb, N., and Fernandez, A. (1999) cdk1- and cdk2-mediated phosphorylation of MyoD Ser200 in growing C2 myoblasts: role in modulating MyoD half-life and myogenic activity, Mol. Cell. Biol., 19, 3167-3176, https://doi.org/10.1128/MCB.19.4.3167.

Supplementary files

Supplementary Files
Action
1. JATS XML
Download
2. Fig. 1. C2C12 cells before differentiation and at different stages of differentiation. Phase-contrast microscopy; 200×. Romanovsky-Giemsa staining of nuclei; L, length of myoblasts; B, width of myoblasts

Download (2MB)
Indexing metadata
3. Fig. 2. Results of western blot analysis of the relative amounts of MyoD (a), MyoG (b), MYH (c), SUCNR1 (d) as a function of the day of differentiation of C2C12 cells. * p < 0.05 - compared to control (before differentiation); ^ p < 0.05 - compared to the first day of differentiation; ˅ p < 0.05 - compared to the fourth day of differentiation

Download (247KB)
Indexing metadata
4. Fig. 3. Results of Western blot analysis of the relative amounts of MyoD and SUCNR1 on days 1 (a) and 4 (b) of differentiation; MYH, MyoG, SUCNR1 - on day 7 of differentiation (c) upon exposure of C2C12 cells to succinic acid at concentrations of 10, 100, 1000 μM. *p < 0.05 - compared to the values of the corresponding day of differentiation without the addition of succinic acid

Download (442KB)
Indexing metadata
5. Fig. 4. Succinate concentration in C2C12 cells during their myogenic differentiation when succinic acid was added to nutrient medium at concentrations of 10 (a), 100 (b) and 1000 (c) μM (detection method - HPLC-MS/MS). * p < 0.05; ** p < 0.01 - statistically significant differences between groups

Download (183KB)
Indexing metadata
6. Fig. 5. Results of Western blot analysis of the relative amounts of MyoD and SUCNR1 on days 1 (a) and 4 (b) of differentiation; MYH, MyoG, SUCNR1 - on day 7 of differentiation (c) when succinic acid at concentrations of 10, 100, 1000 μM, pertussis toxin (Pertussis toxin (PT), 100 ng/ml) and their combined application were applied to C2C12 cells. * p < 0.05 - compared to the values of the corresponding day of differentiation without succinic acid addition

Download (485KB)
Indexing metadata
7. Fig. 6. Inositol monophosphate concentration in C2C12 cells during their myogenic differentiation when succinic acid was added to the nutrient medium at concentrations of 10, 100 and 1000 μM (detection method - HPLC-MS/MS). * p < 0.01 - statistically significant differences with the values of cells before succinate addition, # p < 0.01 - statistically significant differences with the values when succinic acid (100 μM) + pertussis toxin (100 ng/ml) was added

Download (78KB)
Indexing metadata
8. Fig. 7. Putative mechanism of action of succinic acid on the process of myogenic differentiation of C2C12 cells. AC, adenylate cyclase; FLS, phospholipase C; PIF2, phosphatidylinositol-1,5-diphosphate; DAG, diacylglycerol; IF3, inositol-3-phosphate. Mechanism of action via Gαi protein is indicated by bold arrow; mechanism of action via Gγβ protein is indicated by thin arrow; mechanism of action via Ca2+-dependent proteins is indicated by dashed arrow

Download (306KB)
Indexing metadata
9. Fig. 8. C2C12 cells before differentiation (a) and upon stimulation of myogenesis by the addition of 100 μM succinate (b). Phase-contrast microscopy; 200×. Romanovsky-Giemsa staining of nuclei; L - length of myoblasts, B - width of myoblasts

Download (306KB)
Indexing metadata

Copyright (c) 2024 Russian Academy of Sciences