Myogenin Recruits the Histone Chaperone Facilitates Chromatin Transcription (FACT) to Promote Nucleosome Disassembly at Muscle-specific Genes

Alexandra A Lolis,Priya Londhe,Benjamin C Beggs,Stephanie D Byrum,Alan J Tackett,Judith K Davie

doi:10.1074/jbc.m112.426718

Abstract

Facilitates chromatin transcription (FACT) functions to reorganize nucleosomes by acting as a histone chaperone that destabilizes and restores nucleosomal structure. The FACT complex is composed of two subunits: SSRP1 and SPT16. We have discovered that myogenin interacts with the FACT complex. Transfection of FACT subunits with myogenin is highly stimulatory for endogenous muscle gene expression in 10T1/2 cells. We have also found that FACT subunits do not associate with differentiation-specific genes while C2C12 cells are proliferating but are recruited to muscle-specific genes as differentiation initiates and then dissociate as differentiation proceeds. The recruitment is dependent on myogenin, as knockdowns of myogenin show no recruitment of the FACT complex. These data suggest that FACT is involved in the early steps of gene activation through its histone chaperone activities that serve to open the chromatin structure and facilitate transcription. Consistent with this hypothesis, we find that nucleosomes are depleted at muscle-specific promoters upon differentiation and that this activity is dependent on the presence of FACT. Our results show that the FACT complex promotes myogenin-dependent transcription and suggest that FACT plays an important role in the establishment of the appropriate transcription profile in a differentiated muscle cell.

Highlights

The Facilitates chromatin transcription (FACT) complex reorganizes nucleosomes and functions in DNA replication, transcription, and DNA repair
At a 0.1% false discovery rate, 66 proteins were found to coenrich with N-Tandem Affinity Purification (TAP) myogenin, which included putative interacting proteins SSRP1 and SPT16 of the FACT complex
Myogenin-dependent Genes Show an Association of SSRP1 and SPT16 during the Initial Stages of Differentiation—To determine whether FACT was recruited to muscle-specific genes, we assayed for the presence of SPT16 and SSRP1 at two genes that we have shown are dependent on myogenin in vivo, Tnni2 and Leiomodin 2 (Lmod2)

Summary

Background

The FACT complex reorganizes nucleosomes and functions in DNA replication, transcription, and DNA repair. Conclusion: Myogenin recruits the FACT complex to muscle-specific genes where FACT depletes nucleosomes at promoters, facilitating gene expression. The recruitment is dependent on myogenin, as knockdowns of myogenin show no recruitment of the FACT complex These data suggest that FACT is involved in the early steps of gene activation through its histone chaperone activities that serve to open the chromatin structure and facilitate transcription. Consistent with this hypothesis, we find that nucleosomes are depleted at muscle-specific promoters upon differentiation and that this activity is dependent on the presence of FACT. We have discovered that the FACT complex interacts with myogenin and promotes differentiation-specific muscle gene expression. This work aids in our understanding of the factors that control the differentiation program of skeletal muscle and advances our knowledge of the role and recruitment of FACT in skeletal muscle

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION