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Recently, the use of a transgenic mouse model system for Duchenne muscular dystrophy has demonstrated the ability of utrophin to functionally replace dystrophin and alleviate the muscle pathology (see Tinsley, J. M., Potter, A. C., Phelps, S. R., Fisher, R., Trickett, J. I., and Davies, K. E. (1996) Nature 384, 349-353). However, there is currently a clear lack of information concerning the regulatory mechanisms presiding over utrophin expression during normal myogenesis and synaptogenesis. Using in situ hybridization, we show that utrophin mRNAs selectively accumulate within the postsynaptic sarcoplasm of adult muscle fibers. In addition, we demonstrate that a 1.3-kilobase fragment of the human utrophin promoter is sufficient to confer synapse-specific expression to a reporter gene. Deletion of 800 base pairs from this promoter fragment reduces the overall expression of the reporter gene and abolishes its synapse-specific expression. Finally, we also show that utrophin is present at the postsynaptic membrane of ectopic synapses induced to form at sites distant from the original neuromuscular junctions. Taken together, these results indicate that nerve-derived factors regulate locally the transcriptional activation of the utrophin gene in skeletal muscle fibers and that myonuclei located in extrasynaptic regions are capable of expressing utrophin upon receiving appropriate neuronal cues.

Utrophin is a ubiquitously expressed cytoskeletal protein which is an important structural component of the mammalian neuromuscular junction. It shows extensive sequence similarity to dystrophin leading to postulation that utrophin may be able to compensate for the absence of dystrophin in Duchenne muscular dystrophy (DMD) patients. In order to study the transcriptional control of utrophin expression including its regulation at the neuromuscular junction, and as a first step in the development of a potential DMD therapy, we have cloned the utrophin promoter region from human and mouse. The utrophin promoter is associated with a CpG island at the 5'-end of the gene, and sequence analysis of the 5'-UTR reveals several Sp1 binding sites and the absence of TATA or CAAT motifs. Transcription is initiated at one major and three minor sites. Using deletion constructs, we have defined an active promoter region of 155 bp. The first exon and 900 bp upstream display limited sequence conservation between human and mouse. The core sequence TTCCGG of the N box which regulates synaptic expression of other genes is also present and may be involved in regulating the specific expression of utrophin at the postsynaptic membrane. This study provides the basis for the understanding of the regulatory mechanism that controls utrophin expression and provides the data needed to develop methods for the upregulation of utrophin in DMD patients.