Dynamic membrane proteome of adipogenic and myogenic precursors in skeletal muscle highlights EPHA2 may promote myogenic differentiation through ERK signaling

Abstract

The balance of myogenic and adipogenic differentiation is crucial for skeletal muscle homeostasis. Given the vital role of membrane proteins (MBPs) in cell signal perception, membrane proteomics was conducted to delineate mechanisms regulating differentiation of adipogenic and myogenic precursors in skeletal muscle. Adipogenic and myogenic precursors with divergent differentiation potential were isolated from the longissimus dorsi muscle of neonatal pigs by the preplate method. A total of 85 differentially expressed MBPs ( P < 0.05 and fold change ≥1.2 or ≤0.83) between 2 precursors were detected via isobaric tags for relative and absolute quantitation (iTRAQ) assay, including 67 up-regulated and 18 down-regulated in myogenic precursors. Functional enrichment analysis uncovered that myogenic and adipogenic precursors showed significant differences in cytoskeleton organization, syncytium formation, environmental information processing, and organismal systems. Furthermore, key MBPs in regulating cell differentiation were also characterized, including ITGB3, ITGAV, ITPR3, and EPHA2. Noteworthily, EPHA2 was required for myogenic differentiation, and it may promote myogenic differentiation through ERK signaling. Collectively, our study provided an insight into the distinct MBP profile between myogenic and adipogenic precursors in skeletal muscle and served as a solid basis for supporting the role of MBPs in regulating differentiation.—Zhang, X., Wang, L., Qiu, K., Xu, D., Yin, J. Dynamic membrane proteome of adipogenic and myogenic precursors in skeletal muscle highlights EPHA2 may promote myogenic differentiation through ERK signaling.

Global DNA methylation pattern involved in the modulation of differentiation potential of adipogenic and myogenic precursors in skeletal muscle of pigs

“…KEGG analysis based on 153 co-different genes depicted putative regulatory pathways belonging to the cellular process and extracellular environmental processing, through which DNA methylation regulated the distinct differentiation potential of myogenic and adipogenic precursors. These findings were in consistent with our previous study [ 27 ]. Among these pathways, focal adhesion pathway, acting as a bridge between integrin-ECM connection and cytoskeleton [ 52 ], was demonstrated to be largely affected by DNA methylation status [ 53 ].…”

“…Due to the lack of antibodies being used in immunofluorescence histochemistry assay for pigs, we confirmed the identification of these two population cells by their different mRNA expression levels of adipocyte-specific genes and myogenic regulatory factors and their distinct differentiation capacity upon adipogenic and myogenic induction (as shown in Figure S1). In addition, in our previous studies, we have evidenced the high purity of these two subpopulation cells [ 27 , 28 ], and observed significantly higher mRNA expression levels of Desmin and Pax7 in myogenic precursors compared with adipogenic precursors [ 28 ]. Based on the significantly divergent differentiation potential, we can reasonably confirm that these two subpopulation cells are adipogenic and myogenic precursors.…”

“…This study was approved by the Institutional Animal Care and Use Committee (CAU20161110-2) of China Agricultural University. Adipogenic and myogenic precursors were isolated from the skeletal muscle of neonatal pigs using a preplate technique as described previously [ 27 , 28 ]. Briefly, skeletal muscle was minced and digested by 0.17% protease (Sigma-Aldrich, USA) at 37 °C for 1 h, and 0.15% collagenase-type XI solution (Sigma-Aldrich, USA) for 1 h in turn.…”

“…Total RNA was extracted from cells using RNAiso Plus (Takara, China) according to the manufacturer’s instruction. qRT-PCR assays were conducted as previously described [ 27 ]. GAPDH was used as an internal control.…”

Association Analysis of Single-Cell RNA Sequencing and Proteomics Reveals a Vital Role of Ca2+ Signaling in the Determination of Skeletal Muscle Development Potential

“…Besides, the global proteomic profiles of Myo-lineage cells revealed that major differences between obese-and lean-type pigs belong to categories that include muscle development, ion homeostasis, apoptosis, and the MAPK signaling pathway. In a previous study, we showed that the MAPK signaling pathway participates in the mediation of myogenic differentiation [34,35]. Thereby, in the present study, we mainly focused on the proteomic landscape among cell proliferation and differentiation, cellular ion homeostasis, and muscle development.…”