An E box comprises a positional sensor for regional differences in skeletal muscle gene expression and methylation

To dissect the molecular mechanisms conferring positional information in sk eletal muscles, we characterized the control elements responsible for the p ositionally restricted expression patterns of a muscle-specific transgene r eporter, driven by regulatory sequences from the MLC1/3 locus. These sequen ces have previously been shown to generate graded transgene expression in t he segmented axial muscles and their myotomal precursors, fortuitously mark ing their positional address. An evolutionarily conserved E box in the MLC enhancer core, not recognized by MyoD, is a target for a nuclear protein co mplex, present in a variety of tissues, which includes Hox proteins and Zbu 1, a DNA-binding member of the SW12/SNF2 gene family. Mutation of this E bo x in the MLC enhancer has only a modest positive effect on linked CAT gene expression in transfected muscle cells, but when introduced into transgenic mice the same mutation elevates CAT transgene expression in skeletal muscl es, specifically releasing the rostral restriction on MLC-CAT transgene exp ression in the segmented axial musculature. Increased transgene activity re sulting from the E box mutation in the MLC enhancer correlates with reduced DNA methylation of the distal transgenic MLC1 promoter as well as in the e nhancer itself. These results identify an E box and the proteins that bind to it as a positional sensor responsible for regional differences in axial skeletal muscle gene expression and accessibility. (C) 1999 Academic Press.