Muscle Differentiation Is Antagonized by SOX15, a New Member of the SOX Protein Family

Florence Béranger,Catherine Méjean,Brigitte Moniot,Philippe Berta,Marie Vandromme

doi:10.1074/jbc.275.21.16103

Florence Béranger, Catherine Méjean + Show 3 more

Open Access

https://doi.org/10.1074/jbc.275.21.16103

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Abstract

SOX proteins belong to a multigenic family characterized by a unique DNA binding domain, known as the high mobility group box, that is related to that of the testis determining gene SRY. cDNA sequences for more than 30 SOX genes have been identified, and some are known to have diverse roles in vertebrate differentiation and development. Here, we report the isolation and characterization of mouse Sox15 that was uncovered during a screen for high mobility group box containing transcription factors that are expressed at different levels during skeletal muscle differentiation. Sox15 cDNAs were found at a much higher frequency in myoblasts prior to their differentiation into myotubes. Electrophoretic mobility shift assays indicated that recombinant SOX15 protein was capable of binding to a consensus DNA binding site for SOX proteins. When overexpressed in C2C12 myoblasts, wild type SOX15, but not a C-terminal truncated form or the related protein SOX11, specifically inhibited activation of muscle-specific genes and expression of the basic helix-loop-helix myogenic factors myogenin and MyoD, resulting in a failure of the cells to differentiate into myotubes. These results suggest a specific and repressive role for SOX15, requiring the C-terminal domain, during myogenesis.

Highlights

The sex determination gene, SRY, encodes a protein with a 79-amino acid DNA binding motif known as the high mobility group (HMG)1 box [1]
Cloning and Characterization of Sox15—In order to identify Sox genes involved in muscle differentiation, we used a RTPCR screen to search for genes that are differentially expressed between murine C2C12 myoblasts and myotubes
Analysis of the clones differentially expressed between the two cell stages, 18 clones (9% of all clones) corresponding to the Sox15 HMG box were detected in proliferating myoblasts whereas only 2 (1% of all clones) were isolated from differentiated C2C12 cells

Summary

Introduction

The sex determination gene, SRY, encodes a protein with a 79-amino acid DNA binding motif known as the high mobility group (HMG) box [1]. SOX proteins can act as transcription factors and architectural components of chromatin They bind in vitro to the same DNA consensus sequence as SRY ((A/T)(A/T)CAA(A/T)G) [4, 5]. Muscle differentiation is characterized by withdrawal of myoblasts from the cell cycle, induction of muscle-specific gene expression, and cell fusion into multinucleated myotubes. All of these events are coordinated by members of the MyoD family of myogenic bHLH proteins (MyoD, Myf, myogenin, and MRF4) To determine whether Sox genes were involved in skeletal muscle differentiation, we isolated Sox genes that are expressed at different levels in proliferating murine C2C12 myoblasts or fully differentiated myotubes. Myoblasts overexpressing SOX15, under conditions that normally lead to differentiation, (i) were not able to express the early differentiation marker myogenin, (ii) failed to activate muscle-specific gene expression as assessed by a MCK luciferase assay, and (iii) did not fuse into multinu-

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