MYSM1 regulates the proliferation and differentiation of bovine skeletal muscle satellite cells via BRG1-mediated activation of the AKT/mTOR/NF-κB signaling pathway

다국어 초록 (Multilingual Abstract)

Objective: This study investigates the molecular mechanisms by which the deubiquitinase <i>MYSM1</i> regulates the proliferation and differentiation of bovine skeletal muscle satellite cells (BSMSCs), thereby providing new theoretical insights into the regulation of muscle growth and development in beef cattle.Methods: An in vitro model of BSMSCs was established to investigate the role of <i>MYSM1</i>. The expression pattern of <i>MYSM1</i> during cell proliferation and differentiation was analyzed using quantitative real-time polymerase chain reaction (qRT-PCR). <i>MYSM1</i> knockdown models were generated, and the expression of proliferation markers <i>PAX7</i> and <i>Ki67</i>, as well as differentiation markers <i>MYHC</i> and <i>MYOG</i>, were examined by qRT-PCR and Western blotting. Analysis of protein–protein interactions identified <i>BRG1</i> as a potential <i>MYSM1</i> interactor, and its function was subsequently evaluated. Downstream signaling activity was evaluated by examining phosphorylation changes in key components of the AKT/mTOR pathway. In addition, global histone ubiquitination <i>H2AK119ub1</i> and the histone methylation markers <i>H3K4me3</i> and <i>H3K27me3</i> were analyzed following <i>MYSM1</i> knockdown.Results: <i>MYSM1</i> expression was dynamically regulated, exhibiting significant upregulation during differentiation, reaching its highest at days 2–3 (p<0.05). Silencing of <i>MYSM1</i> significantly (p<0.05) decreased the expression of <i>PAX7</i>, <i>Ki67</i>, <i>MYHC</i>, and <i>MYOG</i>. Histone modification analyses demonstrated elevated levels of <i>H2AK119ub1</i> and <i>H3K27me3</i>, along with reduced <i>H3K4me3</i> (p<0.05). Mechanistic studies showed that MYSM1 knockdown significantly reduced <i>BRG1</i> expression (p<0.05), and <i>BRG1</i> silencing similarly decreased proliferation and differentiation markers. Moreover, interference with either <i>MYSM1</i> or <i>BRG1</i> significantly attenuated the activation of the <i>AKT/mTOR/NF-κB</i> signaling pathway, as evidenced by decreased phosphorylation of <i>AKT, mTOR, and p65</i> (p<0.05).Conclusion: <i>MYSM1</i> promotes the proliferation and differentiation of BSMSCs through <i>BRG1</i>-mediated epigenetic regulation and activation of the <i>AKT/mTOR/NF-κB</i> signaling cascade. These findings establish a dual-target framework that advances the understanding of muscle development in beef cattle and offers potential strategies for regenerative therapies.

다국어 초록 (Multilingual Abstract)

Objective: This study investigates the molecular mechanisms by which the deubiquitinase MYSM1 regulates the proliferation and differentiation of bovine skeletal muscle satellite cells (BSMSCs), thereby providing new theoretical insights into the regulation of muscle growth and development in beef cattle.
Methods: An in vitro model of BSMSCs was established to investigate the role of MYSM1.
The expression pattern of MYSM1 during cell proliferation and differentiation was ana lyzed using quantitative real-time polymerase chain reaction (qRT-PCR). MYSM1 knock down models were generated, and the expression of proliferation markers PAX7 and Ki67, as well as differentiation markers MYHC and MYOG, were examined by qRT-PCR and Western blotting. Analysis of protein–protein interactions identified BRG1 as a potential MYSM1 interactor, and its function was subsequently evaluated. Downstream signaling activity was evaluated by examining phosphorylation changes in key components of the AKT/mTOR pathway. In addition, global histone ubiquitination H2AK119ub1 and the histone methylation markers H3K4me3 and H3K27me3 were analyzed following MYSM1 knockdown.
Results: MYSM1 expression was dynamically regulated, exhibiting significant upregula tion during differentiation, reaching its highest at days 2–3 (p<0.05). Silencing of MYSM1 significantly (p<0.05) decreased the expression of PAX7, Ki67, MYHC, and MYOG. His tone modification analyses demonstrated elevated levels of H2AK119ub1 and H3K27me3, along with reduced H3K4me3 (p<0.05). Mechanistic studies showed that MYSM1 knock down significantly reduced BRG1 expression (p<0.05), and BRG1 silencing similarly de creased proliferation and differentiation markers. Moreover, interference with either MYSM1 or BRG1 significantly attenuated the activation of the AKT/mTOR/NF-κB signal ing pathway, as evidenced by decreased phosphorylation of AKT, mTOR, and p65 (p<0.05).
Conclusion: MYSM1 promotes the proliferation and differentiation of BSMSCs through BRG1-mediated epigenetic regulation and activation of the AKT/mTOR/NF-κB signaling cascade. These findings establish a dual-target framework that advances the understanding of muscle development in beef cattle and offers potential strategies for regenerative thera pies.