Int J Biol Sci 2019; 15(4):838-846. doi:10.7150/ijbs.31371
The LIM-Only Protein FHL2 is involved in Autophagy to Regulate the Development of Skeletal Muscle Cell
1. Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu, Sichuan 611130, PR China
2. Animal Breeding and Genetics key Laboratory of Sichuan Province, Sichuan Animal Science Academy, Chengdu, Sichuan, 610066, PR China
* These authors contributed equally to this work.
Scope: Four and a half LIM domain protein 2 (FHL2) is a LIM domain protein expressed in muscle tissue whose deletion is causative of myopathies. Although FHL2 has a confirmed important role in muscle development, its autophagy-related function in muscle differentiation has not been fully determined.
Methods: C2C12 cells were treated with FHL2-konwdown or FHL2-overexpression. The morphology of C2C12 cells was observed by transmission electron microscopy. The mRNA and protein abundances of muscle related genes and autophagy related genes were measured by RT-PCR and western blot. Immunofluorescence and co-immunoprecipitation assay were used to verify the interaction between FHL2 and LC3 protein.
Results: FHL2 silencing reduced LC3-Ⅱ protein expression and the amount of LC3 that co-immunoprecipitated with FHL2, indicating that FHL2 interacts with LC3-Ⅱ in the formation of autophagosomes. Moreover, the expression of muscle development marker genes such as MyoD1 and MyoG was lower in FHL2-silenced C2C12 cells but not in FHL2-overexpressing C2C12 cells. Electron microscopy analysis revealed large empty autophagosomes in FHL2-silenced myoblasts, while flow cytometry suggested that FHL2 silencing made cells more vulnerable to staurosporine-induced cell death.
Conclusion: These results suggest that FHL2 interacts with LC3-Ⅱ in autophagosome formation to regulate the development of muscle cells.
Keywords: FHL2, LIM-domain, autophagy, cell development, C2C12 cells
Liu Z, Han S, Wang Y, Cui C, Zhu Q, Jiang X, Yang C, Du H, Yu C, Li Q, He H, Shen X, Chen Y, Zhang Y, Ye L, Zhang Z, Li D, Zhao X, Yin H. The LIM-Only Protein FHL2 is involved in Autophagy to Regulate the Development of Skeletal Muscle Cell. Int J Biol Sci 2019; 15(4):838-846. doi:10.7150/ijbs.31371. Available from http://www.ijbs.com/v15p0838.htm