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During myogenesis and regeneration, the proliferation and differentiation of myoblasts play key regulatory roles and may be regulated by many genes. In this study, we analyzed the transcriptomic data of chicken primary myoblasts at different periods of proliferation and differentiation with protein‒protein interaction network, and the results indicated that there was an interaction between cyclin-dependent kinase 1 (CDK1) and ribonucleotide reductase regulatory subunit M2 (RRM2). Previous studies in mammals have a role for RRM2in skeletal muscle development as well as cell growth, but the role of RRM2in chicken is unclear. In this study, we investigated the effects of RRM2on skeletal muscle development and regeneration in chickens in vitro and in vivo. The interaction between RRM2and CDK1was initially identified by co-immunoprecipitation and mass spectrometry. Through a dual luciferase reporter assay and quantitative real-time PCR, we identified the core promoter region of RRM2, which is regulated by the SP1transcription factor. In this study, through cell counting kit-8 assays, 5-ethynyl-2′-deoxyuridine incorporation assays, flow cytometry, immunofluorescence staining, and Western blot analysis, we demonstrated that RRM2promoted the proliferation and inhibited the differentiation of myoblasts. In vivo studies showed that RRM2reduced the diameter of muscle fibers and slowed skeletal muscle regeneration. In conclusion, these data provide preliminary insights into the biological functions of RRM2in chicken muscle development and skeletal muscle regeneration.

Skeletal muscle satellite cells are stem cells that are known for their multipotency and ability to proliferate in vitro. However, primary skeletal muscle satellite cells have limited proliferative capacity in vitro, which hinders their study in poultry skeletal muscle. The emergence of immortalization techniques for cells has provided a useful tool to overcome this limitation and explore the functions of skeletal muscle satellite cells. In this study, we achieved the immortalization of chicken skeletal muscle satellite cells by transducing primary cells with TERT (Telomerase reverse transcriptase) amplified from chicken (chTERT) using a lentiviral vector through reconstitution of telomerase activity. The cells successfully bypassed replicative senescence but did not achieve true immortalization. Preliminary functional characterization of the established cell line revealed that the proliferative characteristics and cell cycle profile of the immortalized chicken skeletal muscle satellite cell lines (ICMS) were similar to those of chicken primary muscle satellite cells (CPMSCs). Serum dependency analysis and soft agar assays indicated that ICMS did not undergo malignant transformation. Induced differentiation results demonstrated that ICMS retained their capacity for differentiation. The cell lines established in this study provide an important basis for the establishment of immortalized poultry cell lines and a cell model for the study of poultry skeletal muscle-related functional genes.