Myogenesis pp 25-41 | Cite as

Transdifferentiation of Muscle Satellite Cells to Adipose Cells Using CRISPR/Cas9-Mediated Targeting of MyoD

  • Jingjuan Chen
  • Chao Wang
  • Shihuan KuangEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1889)


Brown adipocytes dissipate energy through non-shivering thermogenesis mediated by UCP1 protein, hence representing a powerful target to overcome obesity due to energy surplus. However, brown adipocytes are scarce in adult humans, especially in obese subjects, urging the development of novel strategies to boost the number of these thermogenic adipocytes from a therapeutical perspective. In this regard, transdifferentiation of myoblasts into brown adipocytes represents a promising approach. Here, we describe a method that we have recently developed to transdifferentiate myoblasts into brown adipocytes through CRISPR/Cas9-medidated targeting of MyoD, the master myogenic regulatory factor.

Key words

Satellite cell Myoblast Adipocyte Transdifferentiation Adipogenesis CRISPR/Cas9 



We thank Jun Wu and Mary Larimore for mouse colony maintenance and members of Kuang laboratory for valuable comments. Jingjuan Chen and Chao Wang have contributed equally to this work.


This work was supported by a grant from the US National Institute of Health (R01AR071649), National Natural Science Foundation of China (81471070) and CAMS Innovation Fund for Medical Sciences (2016-I2M-1-012).


  1. 1.
    Nedergaard J, Bengtsson T, Cannon B (2007) Unexpected evidence for active brown adipose tissue in adult humans. Am J Physiol Endocrinol Metab 293(2):E444–E452. Scholar
  2. 2.
    Cypess AM, Lehman S, Williams G, Tal I, Rodman D, Goldfine AB, Kuo FC, Palmer EL, Tseng Y, Doria A, Kolodny GM, Kahn CR (2009) Identification and importance of brown adipose tissue in adult humans. New Eng J Med 360(15):1509–1517. Scholar
  3. 3.
    Zingaretti MC, Crosta F, Vitali A, Guerrieri M, Frontini A, Cannon B, Nedergaard J, Cinti S (2009) The presence of UCP1 demonstrates that metabolically active adipose tissue in the neck of adult humans truly represents brown adipose tissue. FASEB J 23(9):3113–3120. Scholar
  4. 4.
    Cohade C, Osman M, Pannu HK, Wahl RL (2003) Uptake in supraclavicular area fat ("USA-fat"): description on F-18-FDG PET/CT. J Nucl Med 44(2):170–176PubMedGoogle Scholar
  5. 5.
    Chiba S, Shimada T, Kaneko K, Tomonari K, Satoh Y, Kang M, Fujiwara K, Gotoh K, Masaki T, Katsuragi I, Kakuma T, Hamaguchi K, Yoshimatsu H (2010) Evaluation of human brown adipose tissue using positron emission tomography, computerized tomography and histochemichal studies in association with glucose metabolism. Endocr J 57:S329–S329CrossRefGoogle Scholar
  6. 6.
    Bar-Shalom R, Gaitini D, Keidar Z, Israel O (2004) Non-malignant FDG uptake in infradiaphragmatic adipose tissue: a new site of physiological tracer biodistribution characterised by PET/CT. Eur J Nucl Med Mol Imaging 31(8):1105–1113. Scholar
  7. 7.
    Harms M, Seale P (2013) Brown and beige fat: development, function and therapeutic potential. Nat Med 19(10):1252–1263. Scholar
  8. 8.
    Seale P, Bjork B, Yang WL, Kajimura S, Chin S, Kuang SH, Scime A, Devarakonda S, Conroe HM, Erdjument-Bromage H, Tempst P, Rudnicki MA, Beier DR, Spiegelman BM (2008) PRDM16 controls a brown fat/skeletal muscle switch. Nature 454(7207):961–U927. Scholar
  9. 9.
    Lepper C, Fan CM (2010) Inducible lineage tracing of Pax7-descendant cells reveals embryonic origin of adult satellite cells. Genesis 48(7):424–436. Scholar
  10. 10.
    Lindle RS, Metter EJ, Lynch NA, Fleg JL, Fozard JL, Tobin J, Roy TA, Hurley BF (1997) Age and gender comparisons of muscle strength in 654 women and men aged 20-93 yr. J Appl Physiol 83(5):1581–1587CrossRefGoogle Scholar
  11. 11.
    Farmer SR (2008) Brown fat and skeletal muscle: unlikely cousins? Cell 134(5):726–727. Scholar
  12. 12.
    Rajakumari S, Wu J, Ishibashi J, Lim HW, Giang AH, Won KJ, Reed RR, Seale P (2013) EBF2 determines and maintains brown adipocyte identity. Cell Metab 17(4):562–574. Scholar
  13. 13.
    Nie BM, Nie T, Hui XY, Gu P, Mao LF, Li K, Yuan R, Zheng JS, Wang HX, Tang SB, Zhang Y, Xu T, Xu AM, Wu DH, Ding S (2017) Brown adipogenic reprogramming induced by a small molecule. Cell Rep 18(3):624–635. Scholar
  14. 14.
    Teboul L, Gaillard D, Staccini L, Inadera H, Amri EZ, Grimaldi PA (1995) Thiazolidinediones and fatty-acids convert myogenic cells into adipose-like cells. J Biol Chem 270(47):28183–28187CrossRefGoogle Scholar
  15. 15.
    Fux C, Mitta B, Kramer BP, Fussenegger M (2004) Dual-regulated expression of C/EBP-alpha and BMP-2 enables differential differentiation of C2C12 cells into adipocytes and osteoblasts. Nucleic Acids Res 32(1). Scholar
  16. 16.
    Yu YH, Liu BH, Mersmann HJ, Ding ST (2006) Porcine peroxisome proliferator-activated receptor gamma induces transdifferentiation of myocytes into adipocytes. J Anim Sci 84(10):2655–2665. Scholar
  17. 17.
    Yamanouchi K, Ban A, Shibata S, Hosoyama T, Murakami Y, Nishihara M (2007) Both PPAR gamma and C/EBP alpha are sufficient to induce transdifferentiation of goat fetal myoblasts into adipocytes. J Reprod Dev 53(3):563–572. Scholar
  18. 18.
    Hu ED, Tontonoz P, Spiegelman BM (1995) Transdifferentiation of myoblasts by the adipogenic transcription factors ppar-gamma and C/EBP-alpha. Proc Natl Acad Sci U S A 92(21):9856–9860. Scholar
  19. 19.
    Ohno H, Shinoda K, Ohyama K, Sharp LZ, Kajimura S (2013) EHMT1 controls brown adipose cell fate and thermogenesis through the PRDM16 complex. Nature 504(7478):163. Scholar
  20. 20.
    Sun L, Xie HM, Mori MA, Alexander R, Yuan BB, Hattangadi SM, Liu QQ, Kahn CR, Lodish HF (2011) Mir193b-365 is essential for brown fat differentiation. Nat Cell Biol 13(8):958–U198. Scholar
  21. 21.
    Yin H, Pasut A, Soleimani VD, Bentzinger CF, Antoun G, Thorn S, Seale P, Fernando P, van Ijcken W, Grosveld F, Dekemp RA, Boushel R, Harper ME, Rudnicki MA (2013) MicroRNA-133 controls brown adipose determination in skeletal muscle satellite cells by targeting Prdm16. Cell Metab 17(2):210–224. Scholar
  22. 22.
    Trajkovski M, Ahmed K, Esau CC, Stoffel M (2012) MyomiR-133 regulates brown fat differentiation through Prdm16. Nat Cell Biol 14(12):1330. Scholar
  23. 23.
    Wang C, Liu WY, Nie YH, Qaher M, Horton HE, Yue F, Asakura A, Kuang SH (2017) Loss of MyoD promotes fate transdifferentiation of myoblasts into brown adipocytes. EBioMedicine 16:212–223. Scholar
  24. 24.
    Buckingham M, Relaix F (2015) PAX3 and PAX7 as upstream regulators of myogenesis. Semin Cell Dev Biol 44:115–125. Scholar
  25. 25.
    Christ B, Huang RJ, Scaal M (2007) Amniote somite derivatives. Dev Dyn 236(9):2382–2396. Scholar
  26. 26.
    Kalcheim C (2016) Epithelial-mesenchymal transitions during neural crest and somite development. J Clin Med 5(1). Scholar
  27. 27.
    An YT, Wang G, Diao YR, Long YY, Fu XR, Weng MX, Zhou L, Sun K, Cheung TH, Ip NY, Sun H, Wang HT, Wu ZG (2017) A molecular switch regulating cell fate choice between muscle progenitor cells and brown adipocytes. Dev Cell 41(4):382. Scholar
  28. 28.
    Borensztein M, Viengchareun S, Montarras D, Journot L, Binart N, Lombes M, Dandolo L (2012) Double Myod and Igf2 inactivation promotes brown adipose tissue development by increasing Prdm16 expression. FASEB J 26(11):4584–4591. Scholar
  29. 29.
    Odelberg SJ, Kollhoff A, Keating MT (2000) Dedifferentiation of mammalian myotubes induced by msx1. Cell 103(7):1099–1109. Scholar
  30. 30.
    Yeow K, Phillips B, Dani C, Cabane C, Amri EZ, Derijard B (2001) Inhibition of myogenesis enables adipogenic trans-differentiation in the C2Cl2 myogenic cell line. FEBS Lett 506(2):157–162. Scholar
  31. 31.
    Qi R, Liu H, Wang Q, Wang J, Yang F, Long D, Huang J (2017) Expressions and regulatory effects of P38/ERK/JNK mapks in the adipogenic trans-differentiation of C2C12 myoblasts. Cell Physiol Biochem 44(6):2467–2475. Scholar
  32. 32.
    He Z, Zhu HH, Bauler TJ, Wang J, Ciaraldi T, Alderson N, Li SW, Raquil MA, Ji KH, Wang SF, Shao JH, Henry RR, King PD, Feng GS (2013) Nonreceptor tyrosine phosphatase Shp2 promotes adipogenesis through inhibition of p38 MAP kinase. Proc Natl Acad Sci U S A 110(1):E79–E88. Scholar
  33. 33.
    Sordella R, Jiang W, Chen GC, Curto M, Settleman J (2003) Modulation of rho GTPase signaling regulates a switch between adipogenesis and myogenesis (vol 113, pg 147, 2003). Cell 113(4):547–547. Scholar
  34. 34.
    Ross SE, Hemati N, Longo KA, Bennett CN, Lucas PC, Erickson RL, MacDougald OA (2000) Inhibition of adipogenesis by Wnt signaling. Science 289(5481):950–953. Scholar
  35. 35.
    Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F (2013) Genome engineering using the CRISPR-Cas9 system. Nat Protoc 8(11):2281–2308. Scholar
  36. 36.
    Rudnicki MA, Braun T, Hinuma S, Jaenisch R (1992) Inactivation of myod in mice leads to up-regulation of the myogenic hlh gene MYF-5 and results in apparently normal muscle development. Cell 71(3):383–390. Scholar
  37. 37.
    Motohashi N, Asakura Y, Asakura A (2014) Isolation, culture, and transplantation of muscle satellite cells. J Vis Exp 86.
  38. 38.
    Olson EN, Klein WH (1994) bHLH factors in muscle development: dead lines and commitments, what to leave in and what to leave out. Genes Dev 8(1):1–8CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Animal SciencePurdue UniversityWest LafayetteUSA
  2. 2.Center for Cancer ResearchPurdue UniversityWest LafayetteUSA

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