Analysis of muscle gene transcription in cultured skeletal muscle cells

Abstract

The mechanisms by which muscle gene expression is initiated and maintained are not fully understood. Muscle genes are regulated by combinatorial interactions between numerous transcription factors bound to enhancers and promoters, and their associated protein complexes. Among the most important are the MyoD and MEF2 transcription factor families, but dozens of other factors play important regulatory roles, and many additional transcription factors are certain to be involved. Expression of muscle-specific genes varies among different anatomical muscles and in fast- vs. slow-twitch fiber types, suggesting different mechanisms of regulation in response to diverse physiological cues. Thus, identifying novel transcriptional regulators and interactions is key to understanding how different cells establish the muscle phenotype; it is also critical for developing methods to combat diseases such as muscular dystrophy. Using Muscle creatine kinase as a model, we outline the key steps involved in identifying muscle gene control elements, their binding factors, and mechanisms of transcriptional activation and repression. The basic principles described here can also be applied to the transcriptional analysis of other cell-type specific genes.