CRABP2 Promotes Myoblast Differentiation and Is Modulated by the Transcription Factors MyoD and Sp1 in C2C12 Cells

Jing Yuan,Zhonglin Tang,Shulin Yang,Kui Li,Yao Liang Tang

doi:10.1371/journal.pone.0055479

Jing Yuan, Zhonglin Tang + Show 3 more

Open Access

https://doi.org/10.1371/journal.pone.0055479

Copy DOI

Abstract

Cellular retinoic acid binding protein 2 (CRABP2), a member of a family of specific carrier proteins for Vitamin A, belongs to a family of small cytosolic lipid binding proteins. Our previous study suggested that CRABP2 was involved in skeletal muscle development; however, the molecular function and regulatory mechanism of CRABP2 in myogenesis remained unclear. In this study, we found that the expression of the CRABP2 gene was upregulated during C2C12 differentiation. An over-expression assay revealed that CRABP2 promotes myogenic transformation by regulating the cell cycle during C2C12 differentiation. The region from −459 to −4 bp was identified as the core promoter and contains a TATA box, a GC box and binding sites for the transcription factors MyoD and Sp1. Over-expression, site-directed mutagenesis and EMSA assays indicated that the transcription factors MyoD and Sp1 regulate CRABP2 expression and promote myoblast differentiation in C2C12 cells.

Highlights

Myogenesis can be regarded as a two-step process consisting of the determination, in which the satellite cells commit to the mature muscle lineage, and subsequent differentiation of mononuclear myoblasts to multinuclear myotubes [1]
We found that the CRABP1 and CRBP2 genes were not detected and the mRNA expression of the CRBP1 gene was unchanged, but the Cellular retinoic acid binding protein 2 (CRABP2) gene was markedly upregulated (Figure 1)
The results suggested that CRABP2 was upregulated from day 0 to day 4 during myogenic differentiation in C2C12 cells (Figure 2)

Summary

Introduction

Myogenesis can be regarded as a two-step process consisting of the determination, in which the satellite cells commit to the mature muscle lineage, and subsequent differentiation of mononuclear myoblasts to multinuclear myotubes [1]. A previous study documented that several myogenesis factors (MyoD, Myf, MRF4 and MyoG as well as members of the MyoD gene family) are critical for the determination and terminal differentiation of skeletal muscle cells [2]. In cultured non-muscle cells, the exogenous expression of the MyoD gene family can induce myogenic differentiation [3,4,5,6]. The MyoD gene family proteins bind to the CANNTG sequence ( known as the Ebox sequence), which is present in the promoters and enhancers of many muscle-specific genes [7,8]. Through the DNA-protein interaction, the MyoD gene family induces a conformational change and promotes muscle cell-specific gene transcription. The MyoD gene family promotes cell differentiation and inhibits the cell cycle [9]

Objectives

Methods

Results

Conclusion