Transcriptomic analysis of chicken Myozenin 3 regulation reveals its potential role in cell proliferation

Figures

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  Fig 1. Myoz3 siRNA and overexpression vector screening. (A) Myoz3 expression level in breast muscle, CEFs and Myoblasts. (B) Myoz3 relative expression level in CEFs after transfection with non-specific siRNA (Control), and candidate siRNA. (C) Myoz3 relative expression level in myoblasts after transfection with non-specific siRNA (Control), and candidate siRNA. (D) Myoz3 relative expression level in CEFs after transfection with empty vector and overexpression vector (Over). The error bars represent SEM; all experiments were replicated three times. *P<0.05, **P<0.01, ***P<0.001.

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  Fig 2. The 3rd sample of Inter is abnormal due to inefficient knockdown. (A) An MDS plot of NC and Inter illustrates the abnormality of the 3rd Inter sample. (B) The expression level was not sufficiently knocked down in the 3rd Inter group. The genes that were differentially expressed between the two experiment conditions are presented. The error bars indicate SEM for three replicates (qRT-PCR). Significance was not calculated because to data for each bar do not represent biological replicates.

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  Fig 3. Experimental design and overall results. The expression levels of the top 30 differentially expressed genes are shown in log CPM (copies per million). NC (non-specific control).

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  Fig 4. Validation of differentially expressed genes by qRT-PCR. The relative expression level of data was presented by bar plots with error bars represent SEM. All experiments were replicated three times. (A) MYL9, (B) MYL4 (C) PKM, (D) MPRIP, (E) PDLIM1, (F) MYH10, (G) MYLK2, (H) PPM1J, (I) ECM2, (J) OASL, (K) FAP, (L) CaN, (M) ITGA8. *P<0.05, **P<0.01, ***P<0.001, N.S. means not significant.

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  Fig 5. Enriched biological process (BP) terms. (A) BP terms enriched under the knockdown condition (NC vs Inter). (B) BP terms enriched under the overexpression condition (Control vs Over).

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  Fig 6. Protein interaction networks under the overexpression condition. (A) Biggest network under the overexpression condition, containing 108 genes. (B) Secondary interaction networks of MYH10, containing 47 genes. (C) Network of genes that directly interact with MYH10, containing 11 genes.

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  Fig 7. Proliferation by CCK-8 under both Myoz3 knockdown and overexpression conditions. (A) 24 hours after CEFs transfected with empty vector (Control) or overexpression vector, non-specific siRNA (NC) or Myoz3 siRNA (Inter). (B) 48 hours after CEFs transfected with empty vector (Control) or overexpression vector, non-specific siRNA (NC) or Myoz3 siRNA (Inter). (C) 24 hours after Myoblast transfected with empty vector (Control) or overexpression vector, non-specific siRNA (NC) or Myoz3 siRNA (Inter). (D) 48 hours after myoblasts transfected with empty vector (Control) or overexpression vector, non-specific siRNA (NC) or Myoz3 siRNA (Inter).

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  Fig 8. Myoz3 overexpression results in fast muscle specific gene up-regulated but not slow muscle specific gene. (A) Slow muscle specific gene MYH7B. (B) Fast muscle specific gene MYH1F. NC; non-specific control siRNA. Inter; Myoz3 siRNA. Control; empty vector. Over; Myoz3 overexpression vector. The relative expression level of data was presented by bar plots with error bars represent SEM. All experiments were replicated three times. Error bars represent SEM. *P<0.05, **P<0.01, ***P<0.001.